One Stone Two Birds: Novel Fluorescent Probe ChemistryThat Detects Redox Stress in Cell Culture

Abstract Redox homeostasis is a critical part of physiological processes. Disruption of such homeostasis is associated with various pathological conditions. Tools for studying cellular redox states are important for understanding various biological mechanisms and for the targeted delivery of drugs and/or imaging agents. Along this line, there have been reaction-based fluorescent probes capable of detecting either oxidative stress or hypoxia. However, tools are not available to detect dynamic redox changes in either direction. Toward that end, we describe chemistry that allows for detecting both reduction and oxidation changes using a probe redox pair, moving toward monitoring dynamic redox processes. Embedded in this new approach is also new chemistry for fluorophore activation through the selective reduction of a sulfone group to a thioether under severe hypoxia, complementing the widely used nitro or N-oxide reduction approaches. We demonstrate the feasibility through solution-phase studies and cell-culture work in two cell lines: cancer cells and macrophages. One optimized redox probe pair was studied for its response to both induction of highly reactive oxygen species (hROS) production by turning off the fluorescence and severe hypoxia with fluorescence turn-on. Overall, this approach sets a new direction in designing redox-sensitive tools.