Unsymmetrical Heptamethine Cyanines Prevent Aggregation

Heptamethine cyanine dyes are widely used as near-infrared (NIR) fluorophores in biomedical imaging. However, conventional heptamethine cyanine fluorophores are symmetric in nature which favors fluorophore self-assembly from π–π stacking in aqueous media. This aggregation significantly reduces fluorescence intensity and hampers imaging performance. Although a few non-aggregating heptamethine cyanines have been reported, no general strategy has been established for synthesizing non-aggregating heptamethine cyanine. Here, we introduce a broadly applicable approach to prevent aggregation by breaking the molecular symmetry of heptamethine cyanines through selective substitution along the polymethine backbone. This structural asymmetry disrupts π–π stacking interactions without adversely affecting the desirable photophysical properties of the dyes. The strategy accommodates a wide range of indolenine and polymethine substituents that enable development of a diverse library of non-aggregating heptamethine cyanines. One of the unsymmetrical heptamethine cyanine dyes, SAT-IR-746, was further characterized for its aggregation behavior. SAT-IR-746 remained monomeric in water and across a range of salt concentrations, indicating strong resistance to salt-induced aggregation. Furthermore, a carboxylic acid derivative of SAT-IR-746 was converted to an NHS ester and conjugated to monoclonal antibodies (mAbs) via lysine residues. The resulting IgG–SAT-IR-746 conjugates showed no signs of aggregation, further confirming the dye’s stability in both solution and protein-bound states. Finally, we show the advantages of SAT-IR-746 in live-cell imaging and ex vivo phantom imaging.