Functional Imaging of Oxidative Stress with a Novel PET Imaging Agent, ¹⁸F-5-Fluoro-l-Aminosuberic Acid

Abstract

Glutathione is the predominant endogenous cellular antioxidant, playing a critical role in the cellular defensive response to oxidative stress by neutralizing free radicals and reactive oxygen species. With cysteine as the rate-limiting substrate in glutathione biosynthesis, the cystine/glutamate transporter (system xc−) represents a potentially attractive PET biomarker to enable in vivo quantification of xc− activity in response to oxidative stress associated with disease. We have developed a system xc− substrate that incorporates characteristics of both natural substrates, l-cystine and l-glutamate (l-Glu). l-aminosuberic acid (l-ASu) has been identified as a more efficient system xc− substrate than l-Glu, leading to an assessment of a series of anionic amino acids as prospective PET tracers. Herein, we report the synthesis and in vitro and in vivo validation of a lead candidate, 18F-5-fluoro-aminosuberic acid (18F-FASu), as a PET tracer for functional imaging of a cellular response to oxidative stress with remarkable tumor uptake and retention. Methods 18F-FASu was identified as a potential PET tracer based on an in vitro screening of compounds similar to l-cystine and l-Glu. Affinity toward system xc− was determined via in vitro uptake and inhibition studies using oxidative stress–induced EL4 and SKOV-3 cells. In vivo biodistribution and PET imaging studies were performed in mice bearing xenograft tumors (EL4 and SKOV-3). Results In vitro assay results determined that l-ASu inhibited system xc− as well as or better than l-Glu. The direct comparison of uptake of tritiated compounds demonstrated more efficient system xc− uptake of l-ASu than l-Glu. Radiosynthesis of 18F-FASu allowed the validation of uptake for the fluorine-bearing derivative in vitro. Evaluation in vivo demonstrated primarily renal clearance and uptake of approximately 8 percentage injected dose per gram in SKOV-3 tumors, with tumor-to-blood and tumor-to-muscle ratios of approximately 12 and approximately 28, respectively. 18F-FASu uptake was approximately 5 times greater than 18F-FDG uptake in SKOV-3 tumors. Dynamic PET imaging demonstrated uptake in EL4 tumor xenografts of approximately 6 percentage injected dose per gram and good tumor retention for at least 2 h after injection. Conclusion 18F-FASu is a potentially useful metabolic tracer for PET imaging of a functional cellular response to oxidative stress. 18F-FASu may provide more sensitive detection than 18F-FDG in certain tumors.

In Vivo Imaging of Reactive Oxygen and Nitrogen Species in Murine Colitis

“…Consequent episodes of anesthesia or luciferase administration have not been reported to affect health 58 or learning. 59 Recent developments in the field evaluate the use of nanosensors 60 and RONS-specific PET-tracers 61 for the detection of RONS in inflammation or tumor biology. Although the K8 2/2 colitis and epithelial hyperproliferation occur within the first 2 weeks after birth, 39 our data show that there is a range of RONS produced in young mice while it seems to be stabilized over time in older mice.…”

18F-5-Fluoroaminosuberic Acid as a Potential Tracer to Gauge Oxidative Stress in Breast Cancer Models

“…1) was synthesized on a GE TRACERLab FXFN module using our reported method (17). Radiofluorination was carried out by nucleophilic substitution followed by acidic deprotection.…”

“…We developed a cystine/glutamate analog, 18 F-5-fluoro-aminosuberic acid (FASu), as a potent cystine transporter substrate with the goal of assessing its potential as a diagnostic tracer of OS via x C − activity (17). We previously reported the design, synthesis and in vivo evaluation of 18 F-FASu in EL4 and SKOV3 tumor models (17).…”

“…We previously reported the design, synthesis and in vivo evaluation of 18 F-FASu in EL4 and SKOV3 tumor models (17). Our previous study on 18 F-FASu demonstrated high tumor to background ratios in both tumor types, prompting us to evaluate the specificity of this tracer for xCT and its potential utility in the detection and monitoring of other tumor types.…”

A Continuously Infused Microfluidic Radioassay System for the Characterization of Cellular Pharmacokinetics

“…Thei nvestigation of molecular uptake inside cells is valuable for the understanding of physiologic status (1,2), for the interpretation of molecular imaging results (3,4), and for the development of imaging agents and pharmaceuticals (5). The cellular uptake of a molecule is usually assessed by measuring labeling signals such as radioactivity (2,6), optical signal (7), or magnetic signal (8).…”