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The cell's endogenous antioxidant system is vital to maintain 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

Supplementary Methods and Figures 1 - 6. Fig. S1 shows effect of TBHP treatment on ROS and [18F]FSPG uptake in ovarian cancer cell lines; Fig. S2 illustrates the relationship between glutamate and cystine concentrations on [18F]FSPG uptake; Fig. S3 quantifies [U-13C6, U-15N2]cystine labeling of M+8 GSSG; Fig. S4 indicates extensive disulfide exchange between cystine and NAC; Fig. S5 depicts doxorubicin-mediated alterations in tumor cell redox state and [18F]FSPG; and Fig. S6 illustrates changes in [18F]FSPG and [18F]FDG tumor retention after doxil treatment of mice.

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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.

Supplementary Figures 1 - 6. Fig. S1 shows the ability of PEO1, PEO4 and PEO6 cells to form colonies following cisplatin treatment; Fig. S2 demonstrates that [18F]FDG uptake in A2780 cells is unrelated to drug resistance; Fig. S3 examines the expression of key ABC transporters; Fig. S4 provides representative PET images of mice bearing A2780 WT and A2780 DoxR tumors; Fig. S5 quantifies the [18F]FSPG tumor-to-background ratio in WT and DoxR tumor-bearing mice; and Fig. S6 illustrates [18F]FSPG DoxR tumor retention after doxil treatment of mice.

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.

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.

The cell's endogenous antioxidant system is vital to maintain 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

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.

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.

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 offer immediate practical benefits to access functionalised thiophenols.

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 18F‐fluorination, [18F]4 b was taken up by colorectal tumor cells and trapped through the conversion to its 18F‐labeled carboxylate product under the action of ALDH. In vivo positron emission tomography revealed high uptake of [18F]4 b in the lungs and liver, with radioactivity cleared through the urinary tract. Oxidation of [18F]4 b, however, was observed in vivo, which may limit the tissue penetration of this first‐in‐class radiotracer.

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 HGSOC patients that are refractive to the standard of care. Here, we propose the system x(c)(-) radiotracer (4S)-4-(3-[(18)F]fluoropropyl)-L-glutamate ([(18)F]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 [(18)F]FSPG uptake, both in cells and in animal models of human ovarian cancer. We identified the mechanisms driving differential [(18)F]FSPG cell accumulation and evaluated [(18)F]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 [(18)F]FSPG cell accumulation in drug-resistant versus drug-sensitive cells. Decreased [(18)F]FSPG uptake in drug-resistant cells was a consequence of changes in intracellular cystine, a key precursor in GSH biosynthesis. In vivo, [(18)F]FSPG uptake was decreased ~80% in chemotherapy-resistant A2780 tumors compared to parental drug-sensitive tumors, with non-responding tumors displaying high levels of oxidised-to-reduced GSH. Treatment of drug-resistant A2780 tumors with doxorubicin resulted in no detectable change in tumor volume, GSH or [(18)F]FSPG uptake. CONCLUSIONS: This study demonstrates the ability of [(18)F]FSPG to detect upregulated antioxidant pathways present in drug-resistant cancer. [(18)F]FSPG may therefore enable the identification of HGSOC patients that are refractory to standard-of-care, allowing the transferral of drug-resistant patients to alternative therapies, thereby improving outcomes in this disease.

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.

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 x

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 [ 18 F]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 [ 18 F]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 [ 18 F]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.

Two efficient phase-transfer-catalyzed protocols for the diastereoselective synthesis of cis and trans isomers of 2,6-diaryltetrahydrothiopyran-4-ones (2,6-DATHTPs) have been developed. In a study of the scope of the reactions, differently substituted 2,6-DATHTPs were successfully accessed in high yields and diastereomeric excessses on both experimental and preparative scales.

Introduction: Akin to many cancers, enhanced18F-FDG-PET signal has been associated with fibrotic lesions in pulmonary fibrosis. In tumors, elevated glucose uptake is indicative of a metabolic switch to aerobic glycolysis (AG), critical to fuel biosynthetic demand. De novo expression of the low-activity isoform of the ‘glycolytic gatekeeper’ pyruvate kinase, PKM2, is a key driver of AG and pharmacological modulation of PKM2 is known to attenuate tumor proliferation and tumorigenesis. Aim: We hypothesised that the fibroproliferative response to bleomycin lung injury is characterised by metabolic reprogramming driving an enhanced glucose requirement of cells in the injured lesions. We investigated glucose uptake and modulation of the glycolytic marker PKM2 in the bleomycin mouse model. Methods&Results: Autoradiography of18F-FDG uptake in cryofrozen lung sections at day 28 post-bleomycin showed a significant ›50% increase in18F-FDG uptake in bleomycin-challenged compared to uninjured lungs. The regions of highest18F-FDG uptake corresponded to dense fibrotic regions. Immunohistochemistry revealed that PKM2 was localised to multiple cell types in fibrotic lung lesions, including a-SMA-positive myofibroblasts. A small molecule activator of PKM2, TEPP-46, achieved a significant ~50% increase in PKM2 activity in the bleomycin-injured lung. However, chronic TEPP-46 administration from day 5 post-injury had no significant effect on lung fibrosis quantified by microCt at day 28. Conclusions: We show that glucose uptake is significantly increased in fibrotic lung lesions. Although fibrotic lesions express PKM2, increasing the activity of this isoform was not sufficient to influence progression of fibrosis in bleomycin lung injury

African trypanosomiasis, Chagas disease, and leishmaniasis are human infectious diseases caused by various kinetoplastid parasites. Trypanothione reductase (TR) is a flavoenzyme unique to these parasites that is responsible for maintaining trypanothione (bis(glutathionyl)spermidine) in its reduced dithiol form. This enzyme plays a crucial role in the thiol redox metabolism and is essential in vivo for all trypanosomatids living in the human host studied so far. These findings make the flavoenzyme a promising target for anti-kinetoplastidal drug development. In this chapter, we examine the work published in the field of redox-active agents acting as substrates of the NADPH-dependent TR-based system. We also highlight our own work on trypanothione-reactive agents and discuss how these compounds might be developed as potential specific lead compounds to fight kinetoplastidal parasites.

As a twofold Michael system, the diarylideneacetone core is of particular interest in organic synthesis and for therapeutic applications. To overcome the drawbacks of the classical Claisen–Schmidt protocol, a new methodology for the synthesis of dissymmetric (hetero)diarylideneacetones has been developed. Conditions were optimized with a Box–Behnken design of experiment. The milder reaction conditions allow the efficient preparation of fluorinated, or heteroaromatic, dissymmetric diarylideneacetones which cannot be obtained through the classical Claisen–Schmidt protocol.

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.

Two efficient phase-transfer-catalyzed protocols for the diastereoselective synthesis of cis and trans isomers of 2,6-diaryltetrahydrothiopyran-4-ones (2,6-DATHTPs) are developed.

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.