Method To Diversify Cyanine Chromophore Functionality Enables Improved Biomolecule Tracking and Intracellular Imaging

Area of Science:

• Chemical synthesis
• Biomedical imaging
• Fluorescence microscopy

Background:

• Heptamethine indocyanines are crucial for near-infrared (NIR) imaging but have limited synthetic routes.
• Existing methods for synthesizing these probes are often inefficient or restrictive.

Purpose of the Study:

• To develop a versatile and high-yielding synthetic method for heptamethine indocyanines.
• To create novel NIR probes for improved protein-targeted tumor imaging.
• To engineer cyclizing heptamethine indocyanines for enhanced cellular uptake and imaging properties.

Main Methods:

• Utilized pyridinium benzoxazole (PyBox) salts as novel precursors for heptamethine indocyanine synthesis.
• Employed an iterative approach to optimize probes for protein-targeted tumor imaging.
• Designed cyclizing heptamethine indocyanines by modifying electrophilic and nucleophilic components.
• Demonstrated no-wash live cell imaging with a chloroalkane derivative using HaloTag technology.

Main Results:

• The PyBox method provides a high-yielding and simple route to heptamethine indocyanines with new functionalities.
• Optimized probes showed increased tumor specificity for antibody and nanobody conjugates compared to existing NIR fluorophores.
• Engineered cyclizing heptamethine indocyanines exhibited tunable solvent sensitivity and efficient cellular uptake.
• A specific derivative enabled highly effective no-wash live cell imaging of organelles.

Conclusions:

• The reported chemistry expands accessible chromophore functionality for NIR probes.
• This new synthetic approach enables the development of advanced NIR probes for diverse imaging applications.
• The study highlights improved specificity in tumor imaging and enhanced capabilities in live cell analysis.