Investigation of the ultrasound-mediated toxicity mechanisms of IR780 iodide

IR780 iodide is a lipophilic cation heptamethine dye that has emerged as a potential fluorescent probe for in vivo tumor imaging. Previous work has shown it to be a sono- and photo-sensitizer (sound- and light-activated molecule) suitable for use in sonodynamic therapy (SDT) due to its proposed ability to produce ROS when ‘activated' by ultrasound. This study evaluated IR780 iodide as an SDT agent in vitro using A549s, HeLas, and HeLa S3 cell lines, a broad range of ultrasound and light parameters, multiple ultrasound and light systems, and with and without cavitation nuclei. Through temporal uncoupling of ultrasound application and compound administration in vitro and in vivo, evaluation of cavitation-only related cell death, assessment of the dark toxicity of IR780 iodide, and development of positive controls for cell permeabilization (sonoporation), this study shows that dark toxicity and cavitation play an important role in IR780 SDT-induced cell death. This potentially explains the high levels of cell death observed for comparatively low concentrations of ROS. Additionally, this study proposes a standard set of controls for SDT mechanistic studies, which were used to further help study the mechanisms of other SDT drugs including Rose Bengal, 5-aminolevulinic acid, and indocyanine green.