Abstract
BACKGROUND: Mercury-197m ((197m)Hg, t(1/2) = 23.8 h) and mercury-197g ((197g)Hg, t(1/2) = 64.14 h) possess favorable nuclear properties for imaging and targeted therapy, but the development of suitable chelators for mercury-based radiopharmaceuticals remains underexplored. Additionally, accurate imaging and quantification of mercury isotopes, particularly in dual-isotope formats, require tools that account for their complex decay schemes. Phantom imaging studies are essential for validating spatial resolution, quantitative accuracy, and isotope-specific calibration prior to in vivo application. In this study, we investigated the commercially available ligand H(4)Tetrathiol for chelation of [(197m/g)Hg]Hg(2+) and developed a robust imaging and quantification pipeline to support the use of these nuclear isomers in preclinical imaging. RESULTS: Radiolabeling of H(4)Tetrathiol yielded exceptionally efficient complexation, achieving the lowest ligand-to-metal ratio reported for radio-mercury. The resulting [(197m/g)Hg]Hg(2+)-complex demonstrated high in vitro stability in the presence of serum proteins, glutathione, and competing biologically relevant metal ions, though it exhibited kinetic lability when challenged with excess HgCl₂. In vivo biodistribution studies in mice showed a distinct pharmacokinetic profile from unchelated [(197m/g)Hg]HgCl₂, suggesting in vivo complex stability. Phantom imaging studies with a high sensitivity collimator demonstrated submillimeter resolution (≥ 1.1 mm) for both (197g)Hg and (197m)Hg, with decay behavior consistent with known half-lives. To facilitate accurate quantification, we developed HgQuant, a Python-based tool for isotope-specific calibration, Bateman decay correction, and automated dual-isotope analysis. This tool enabled reproducible, time-resolved quantification in both phantom and in vivo settings. CONCLUSIONS: These results establish Tetrathiol as a promising scaffold for mercury-based theranostics, offering efficient radiolabeling and in vivo stability. The integration of high-resolution imaging and HgQuant-based quantification of each isomer establishes a comprehensive framework for advancing [(197m/g)Hg]Hg radiopharmaceutical development.