Investigation of cyanine dyes for in vivo optical imaging of altered mitochondrial membrane potential in tumors

Mitochondrial membrane potential (Δ ψ m ) alteration is an important target for cancer diagnosis. In this study, we designed a series of near‐infrared fluorescent cationic cyanine dyes with varying alkyl chain lengths ( IC 7‐1 derivatives) to provide diverse lipophilicities and serum albumin‐binding rates, and we evaluated the usefulness of these derivatives for in vivo Δ ψ m imaging. IC 7‐1 derivatives with side chains from methyl to hexyl ( IC 7‐1‐Me to IC 7‐1‐He) were synthesized, and their optical properties were measured. Cellular uptake and intracellular distribution were investigated with depolarized HeLa cells from carbonyl cyanine m ‐chlorophenylhydrazone ( CCCP ) treatment using a spectrofluorometer and a fluorescence microscope. Serum albumin‐binding rates were evaluated using albumin‐binding inhibitors. In vivo optical imaging was performed with HeLa cell xenograft mice following intravenous administration of IC 7‐1 derivatives with or without warfarin and CCCP as in vivo blocking agents. IC 7‐1 derivatives showing maximum excitation and emission wavelengths at 823 nm and ~845 nm, respectively, were synthesized. IC 7‐1‐Me to ‐Bu showed fluorescence in mitochondria that decreased with CCCP treatment in a concentration‐dependent manner, which showed that IC 7‐1‐Me to ‐Bu successfully indicated Δ ψ m . Tumors were clearly visualized after IC 7‐1‐Bu administration. Treatment with warfarin or CCCP significantly decreased IC 7‐1‐Bu fluorescence in the tumor region. In summary, IC 7‐1‐Bu exhibited fluorescence localized to mitochondria dependent on Δ ψ m , which enabled clear in vivo tumor imaging via serum albumin as a drug carrier for effective tumor targeting. Our data suggest that IC 7‐1‐Bu is a promising NIR probe for in vivo imaging of the altered Δ ψ m of tumor cells.