Chemoselective Synthesis and Anti-Kinetoplastidal Properties of 2,6-Diaryl-4H-tetrahydro-thiopyran-4-one S-Oxides: Their Interplay in a Cascade of Redox Reactions from Diarylideneacetones.

2,6-Diaryl-4H-tetrahydro-thiopyran-4-ones and corresponding sulfoxide and sulfone derivatives were designed to lower the major toxicity of their parent anti-kinetoplatidal diarylideneacetones through a prodrug effect. Novel diastereoselective methodologies were developed and generalized from diarylideneacetones and 2,6-diaryl-4H-tetrahydro-thiopyran-4-ones to allow the introduction of a wide substitution profile and to prepare the related S-oxides. The in vitro biological activity and selectivity of diarylideneacetones, 2,6-diaryl-4H-tetrahydro-thiopyran-4-ones, and their S-sulfoxide and sulfone metabolites were evaluated against Trypanosoma brucei brucei, Trypanosoma cruzi, and various Leishmania species in comparison with their cytotoxicity against human fibroblasts hMRC-5. The data revealed that the sulfides, sulfoxides, and sulfones, in which the Michael acceptor sites are temporarily masked, are less toxic against mammal cells while the anti-trypanosomal potency was maintained against T. b. brucei, T. cruzi, L. infantum, and L. donovani, thus confirming the validity of the prodrug strategy. The mechanism of action is proposed to be due to the involvement of diarylideneacetones in cascades of redox reactions involving the trypanothione system. After Michael addition of the dithiol to the double bonds, resulting in an elongated polymer, the latter-upon S-oxidation, followed by syn-eliminations-fragments, under continuous release of reactive oxygen species and sulfenic/sulfonic species, causing the death of the trypanosomal parasites in the micromolar or submicromolar range with high selectivity indexes.

Mapping Aldehyde Dehydrogenase 1A1 Activity using an [18 F]Substrate-Based Approach.

Aldehyde dehydrogenases (ALDHs) catalyze the oxidation of aldehydes to carboxylic acids. Elevated ALDH expression in human cancers is linked to metastases and poor overall survival. Despite ALDH being a poor prognostic factor, the non-invasive assessment of ALDH activity in vivo has not been possible due to a lack of sensitive and translational imaging agents. Presented in this report are the synthesis and biological evaluation of ALDH1A1-selective chemical probes composed of an aromatic aldehyde derived from N,N-diethylamino benzaldehyde (DEAB) linked to a fluorinated pyridine ring either via an amide or amine linkage. Of the focused library of compounds evaluated, N-ethyl-6-(fluoro)-N-(4-formylbenzyl)nicotinamide 4 b was found to have excellent affinity and isozyme selectivity for ALDH1A1 in vitro. Following 18 F-fluorination, [18 F]4 b was taken up by colorectal tumor cells and trapped through the conversion to its 18 F-labeled carboxylate product under the action of ALDH. In vivo positron emission tomography revealed high uptake of [18 F]4 b in the lungs and liver, with radioactivity cleared through the urinary tract. Oxidation of [18 F]4 b, however, was observed in vivo, which may limit the tissue penetration of this first-in-class radiotracer.

Ex vivo characterization of neuroinflammatory and neuroreceptor changes during epileptogenesis using candidate positron emission tomography biomarkers.

OBJECTIVE: Identification of patients at risk of developing epilepsy before the first spontaneous seizure may promote the development of preventive treatment providing opportunity to stop or slow down the disease. METHODS: As development of novel radiotracers and on-site setup of existing radiotracers is highly time-consuming and expensive, we used dual-centre in vitro autoradiography as an approach to characterize the potential of innovative radiotracers in the context of epilepsy development. Using brain slices from the same group of rats, we aimed to characterise the evolution of neuroinflammation and expression of inhibitory and excitatory neuroreceptors during epileptogenesis using translational positron emission tomography (PET) tracers; 18 F-flumazenil (18 F-FMZ; GABAA receptor), 18 F-FPEB (metabotropic glutamate receptor 5; mGluR5), 18 F-flutriciclamide (translocator protein; TSPO, microglia activation) and 18 F-deprenyl (monoamine oxidase B, astroglia activation). Autoradiography images from selected time points after pilocarpine-induced status epilepticus (SE; baseline, 24 and 48 hours, 5, 10 and 15 days and 6 and 12-14 weeks after SE) were normalized to a calibration curve, co-registered to an MRI-based 2D region-of-interest atlas, and activity concentration (Bq/mm2 ) was calculated. RESULTS: In epileptogenesis-associated brain regions, 18 F-FMZ and 18 F-FPEB showed an early decrease after SE. 18 F-FMZ decrease was maintained in the latent phase and further reduced in the chronic epileptic animals, while 18 F-FPEB signal recovered from day 10, reaching baseline levels in chronic epilepsy. 18 F-flutriciclamide showed an increase of activated microglia at 24 hours after SE, peaking at 5-15 days and decreasing during the chronic phase. On the other hand, 18 F-deprenyl autoradiography showed late astrogliosis, peaking in the chronic phase. SIGNIFICANCE: Autoradiography revealed different evolution of the selected targets during epileptogenesis. Our results suggest an advantage of combined imaging of inter-related targets like glutamate and GABAA receptors, or microglia and astrocyte activation, in order to identify important interactions, especially when using PET imaging for the evaluation of novel treatments.

Ring-Closing Synthesis of Dibenzothiophene Sulfonium Salts and Their Use as Leaving Groups for Aromatic 18F-Fluorination.

Herein, we report a novel intramolecular ring-closing reaction of biaryl thioethers that give access to highly functionalized dibenzothiophene sulfonium salts under mild conditions. The resulting precursors react regioselectively with [18F]fluoride to give [18F]fluoroarenes in predictable radiochemical yields. The strategy expands the available radiochemical space and provides superior labeling efficiency for clinically relevant PET tracers.

Characterization of tau positron emission tomography tracer [18F]AV-1451 binding to postmortem tissue in Alzheimer's disease, primary tauopathies, and other dementias.

INTRODUCTION: Aggregation of tau is a hallmark of many neurodegenerative diseases, and tau imaging with positron emission tomography (PET) may allow early diagnosis and treatment monitoring. We assessed binding of the PET tracer [18F]AV-1451 in a range of dementias. METHODS: Phosphorimaging was used to quantify binding to postmortem brain tissue from 33 patients with different, histopathologically characterized, neurodegenerative dementias. RESULTS: [18F]AV-1451 showed high specific binding in cases with Alzheimer's disease (AD), moderate binding in Pick's disease and frontotemporal dementia with parkinsonism-17, and low but displaceable binding in corticobasal degeneration, progressive supranuclear palsy, non-tau proteinopathies, and in controls without pathology. Tracer binding did not correlate with tau load within disease groups. DISCUSSION: [18F]AV-1451 binds to tau in AD, and some other tauopathies. However, evidence for a non-tau binding site and lack of correlation between tracer binding and antibody staining suggest that reliable quantification of tau load with this tracer is problematic.

In Silico Mining for Antimalarial Structure-Activity Knowledge and Discovery of Novel Antimalarial Curcuminoids.

Malaria is a parasitic tropical disease that kills around 600,000 patients every year. The emergence of resistant Plasmodium falciparum parasites to artemisinin-based combination therapies (ACTs) represents a significant public health threat, indicating the urgent need for new effective compounds to reverse ACT resistance and cure the disease. For this, extensive curation and homogenization of experimental anti-Plasmodium screening data from both in-house and ChEMBL sources were conducted. As a result, a coherent strategy was established that allowed compiling coherent training sets that associate compound structures to the respective antimalarial activity measurements. Seventeen of these training sets led to the successful generation of classification models discriminating whether a compound has a significant probability to be active under the specific conditions of the antimalarial test associated with each set. These models were used in consensus prediction of the most likely active from a series of curcuminoids available in-house. Positive predictions together with a few predicted as inactive were then submitted to experimental in vitro antimalarial testing. A large majority from predicted compounds showed antimalarial activity, but not those predicted as inactive, thus experimentally validating the in silico screening approach. The herein proposed consensus machine learning approach showed its potential to reduce the cost and duration of antimalarial drug discovery.

Correction: Pacelli et al. Fully Automated, High-Dose Radiosynthesis of [18F]PARPi. Pharmaceuticals 2022, 15, 865.

Corentin Warnier, Thibault Gendron, Muhammad Otabashi, Charles Vriamont and Alex Jackson were not included as authors in the original publication [...].

multi-patient dose synthesis of [18F]Flumazenil via a copper-mediated 18F-fluorination.

BACKGROUND: Flumazenil (FMZ) is a functionally silent imidazobenzodiazepine which binds to the benzodiazepine binding site of approximately 75% of the brain γ-aminobutyric acid-A receptors (GABAARs). Positron Emission Tomography (PET) imaging of the GABAARs with [11C]FMZ has been used to evidence alterations in neuronal density, to assess target engagement of novel pharmacological agents, and to study disorders such as epilepsy and Huntington's disease. Despite the potential of FMZ PET imaging the short half-life (t1/2) of carbon-11 (20 min) has limited the more widespread clinical use of [11C]FMZ. The fluorine-18 (18F) isotopologue with a longer t1/2 (110 min) is ideally suited to address this drawback. However, the majority of current radiochemical methods for the synthesis of [18F]FMZ are non-trivial and low yielding. We report a robust, automated protocol that is good manufacturing practice (GMP) compatible, and yields multi-patient doses of [18F]FMZ. RESULTS: The fully automated synthesis was developed on the Trasis AllinOne (AIO) platform using a single-use cassette. [18F]FMZ was synthesized in a one-step procedure from [18F]fluoride, via a copper-mediated 18F-fluorination of a boronate ester precursor. Purification was performed by semi-preparative radio-HPLC and the collected fraction formulated directly into the final product vial. The overall process from start of synthesis to delivery of product is approximately 55 min. Starting with an initial activity of 23.6 ± 5.8 GBq (n = 3) activity yields of [18F]FMZ were 8.0 ± 1 GBq (n = 3). The synthesis was successfully reproduced at two independent sites, where the product passed quality control release criteria in line with the European Pharmacopoeia standards and ICH Q3D(R1) guidelines to be suitable for human use. CONCLUSION: Reported is a fully automated cassette-based synthesis of [18F]FMZ that is Good Manufacturing Practice (GMP) compatible and produces multi-patient doses of [18F]FMZ.

Development of [18F]AldoView as the First Highly Selective Aldosterone Synthase PET Tracer for Imaging of Primary Hyperaldosteronism.

The purpose of this study was to synthesize a fluorine-18 labeled, highly selective aldosterone synthase (hCYP11B2) inhibitor, [18F]AldoView, and to assess its potential for the detection of aldosterone-producing adenomas (APAs) with positron emission tomography in patients with primary hyperaldosteronism (PHA). Using dibenzothiophene sulfonium salt chemistry, [18F]AldoView was obtained in high radiochemical yield in one step from [18F]fluoride. In mice, the tracer showed a favorable pharmacokinetic profile, including rapid distribution and clearance. Imaging in the adrenal tissue from patients with PHA revealed diffuse binding patterns in the adrenal cortex, avid binding in some adenomas, and "hot spots" consistent with aldosterone-producing cell clusters. The binding pattern was in good visual agreement with the antibody staining of hCYP11B2 and distinguished areas with normal and excessive hCYP11B2 expression. Taken together, [18F]AldoView is a promising tracer for the detection of APAs in patients with PHA.

Iron(II)/Persulfate Mediated Newman-Kwart Rearrangement.

Herein, we report that iron(II)/ammonium persulfate in aqueous acetonitrile mediates the Newman-Kwart rearrangement of O-aryl carbamothioates. Electron-rich substrates react rapidly under moderate heating to afford the rearranged products in excellent yields. The mild conditions, rapid reaction rates, and suitability for scale up offers immediate practical benefits to access functionalized thiophenols.

One-Pot Radiosynthesis and Biological Evaluation of a Caspase-3 Selective 5-[123,125I]iodo-1,2,3-triazole derived Isatin SPECT Tracer.

Induction of apoptosis is often necessary for successful cancer therapy, and the non-invasive monitoring of apoptosis post-therapy could assist in clinical decision making. Isatins are a class of compounds that target activated caspase-3 during apoptosis. Here we report the synthesis of the 5-iodo-1,2,3-triazole (FITI) analog of the PET tracer [18F]ICMT11 as a candidate tracer for imaging of apoptosis with SPECT, as well as PET. Labelling with radioiodine (123,125I) was achieved in 55 ± 12% radiochemical yield through a chelator-accelerated one-pot cycloaddition reaction mediated by copper(I) catalysis. The caspase-3 binding affinity and selectivity of FITI compares favourably to that of [18F]ICMT11 (Ki = 6.1 ± 0.9 nM and 12.4 ± 4.7 nM, respectively). In biodistribution studies, etoposide-induced cell death in a SW1222 xenograft model resulted in a 2-fold increase in tumour uptake of the tracer. However, the tumour uptake was too low to allow in vivo imaging of apoptosis with SPECT.

Measurement of Tumor Antioxidant Capacity and Prediction of Chemotherapy Resistance in Preclinical Models of Ovarian Cancer by Positron Emission Tomography.

PURPOSE: Drug resistance is a major obstacle for the effective treatment of patients with high-grade serous ovarian cancer (HGSOC). Currently, there is no satisfactory way to identify patients with HGSOC that are refractive to the standard of care. Here, we propose the system xc - radiotracer (4S)-4-(3-[18F]fluoropropyl)-l-glutamate ([18F]FSPG) as a non-invasive method to measure upregulated antioxidant pathways present in drug-resistant HGSOC. EXPERIMENTAL DESIGN: Using matched chemotherapy sensitive and resistant ovarian cancer cell lines, we assessed their antioxidant capacity and its relation to [18F]FSPG uptake, both in cells and in animal models of human ovarian cancer. We identified the mechanisms driving differential [18F]FSPG cell accumulation and evaluated [18F]FSPG tumor uptake as predictive marker of treatment response in drug-resistant tumors. RESULTS: High intracellular glutathione (GSH) and low reactive oxygen species corresponded to decreased [18F]FSPG cell accumulation in drug-resistant versus drug-sensitive cells. Decreased [18F]FSPG uptake in drug-resistant cells was a consequence of changes in intracellular cystine, a key precursor in GSH biosynthesis. In vivo, [18F]FSPG uptake was decreased nearly 80% in chemotherapy-resistant A2780 tumors compared with parental drug-sensitive tumors, with nonresponding tumors displaying high levels of oxidized-to-reduced GSH. Treatment of drug-resistant A2780 tumors with doxorubicin resulted in no detectable change in tumor volume, GSH, or [18F]FSPG uptake. CONCLUSIONS: This study demonstrates the ability of [18F]FSPG to detect upregulated antioxidant pathways present in drug-resistant cancer. [18F]FSPG may therefore enable the identification of patients with HGSOC that are refractory to standard of care, allowing the transferal of drug-resistant patients to alternative therapies, thereby improving outcomes in this disease.

Assessment of Tumor Redox Status through (S)-4-(3-[18F]fluoropropyl)-L-Glutamic Acid PET Imaging of System xc - Activity.

The cell's endogenous antioxidant system is vital to maintenance of redox homeostasis. Despite its central role in normal and pathophysiology, no noninvasive tools exist to measure this system in patients. The cystine/glutamate antiporter system xc - maintains the balance between intracellular reactive oxygen species and antioxidant production through the provision of cystine, a key precursor in glutathione biosynthesis. Here, we show that tumor cell retention of a system xc --specific PET radiotracer, (S)-4-(3-[18F]fluoropropyl)-L-glutamic acid ([18F]FSPG), decreases in proportion to levels of oxidative stress following treatment with a range of redox-active compounds. The decrease in [18F]FSPG retention correlated with a depletion of intracellular cystine resulting from increased de novo glutathione biosynthesis, shown through [U-13C6, U-15N2]cystine isotopic tracing. In vivo, treatment with the chemotherapeutic doxorubicin decreased [18F]FSPG tumor uptake in a mouse model of ovarian cancer, coinciding with markers of oxidative stress but preceding tumor shrinkage and decreased glucose utilization. Having already been used in pilot clinical trials, [18F]FSPG PET could be rapidly translated to the clinic as an early redox indicator of tumor response to treatment. SIGNIFICANCE: [18F]FSPG PET imaging provides a sensitive noninvasive measure of tumor redox status and provides an early marker of tumor response to therapy.See related commentary by Lee et al., p. 701.

Sulfonium salts as leaving groups for aromatic labelling of drug-like small molecules with fluorine-18.

Positron emission tomography (PET) is unique in that it allows quantification of biochemical processes in vivo, but difficulties with preparing suitably labelled radiotracers limit its scientific and diagnostic applications. Aromatic [(18)F]fluorination of drug-like small molecules is particularly challenging as their functional group compositions often impair the labelling efficiency. Herein, we report a new strategy for incorporation of (18)F into highly functionalized aromatic compounds using sulfonium salts as leaving groups. The method is compatible with pharmacologically relevant functional groups, including aliphatic amines and basic heterocycles. Activated substrates react with [(18)F]fluoride at room temperature, and with heating the reaction proceeds in the presence of hydrogen bond donors. Furthermore, the use of electron rich spectator ligands allows efficient and regioselective [(18)F]fluorination of non-activated aromatic moieties. The method provides a broadly applicable route for (18)F labelling of biologically active small molecules, and offers immediate practical benefits for drug discovery and imaging with PET.

Development of Fluorine-18 Labeled Metabolically Activated Tracers for Imaging of Drug Efflux Transporters with Positron Emission Tomography.

Increased activity of efflux transporters, e.g., P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP), at the blood-brain barrier is a pathological hallmark of many neurological diseases, and the resulting multiple drug resistance represents a major clinical challenge. Noninvasive imaging of transporter activity can help to clarify the underlying mechanisms of drug resistance and facilitate diagnosis, patient stratification, and treatment monitoring. We have developed a metabolically activated radiotracer for functional imaging of P-gp/BCRP activity with positron emission tomography (PET). In preclinical studies, the tracer showed excellent initial brain uptake and clean conversion to the desired metabolite, although at a sluggish rate. Blocking with P-gp/BCRP modulators led to increased levels of brain radioactivity; however, dynamic PET did not show differential clearance rates between treatment and control groups. Our results provide proof-of-concept for development of prodrug tracers for imaging of P-gp/BCRP function in vivo but also highlight some challenges associated with this strategy.