Doubly Strapped Zwitterionic NIR‐I and NIR‐II Heptamethine Cyanine Dyes for Bioconjugation and Fluorescence Imaging

Abstract

Heptamethine cyanine dyes enable deep tissue fluorescence imaging in the near infrared (NIR) window. Small molecule conjugates of the benchmark dye ZW800-1 have been tested in humans. However, longterm imaging protocols using ZW800-1 conjugates are limited by their instability, primarily because the chemically labile C4'-O-aryl linker is susceptible to cleavage by biological nucleophiles. Here, we report a modular synthetic method that produces novel doubly strapped zwitterionic heptamethine cyanine dyes, including a structural analogue of ZW800-1, with greatly enhanced dye stability. NIR-I and NIR-II versions of these doubly strapped dyes can be conjugated to proteins, including monoclonal antibodies, without causing undesired fluorophore degradation or dye stacking on the protein surface. The fluorescent antibody conjugates show excellent tumor-targeting specificity in a xenograft mouse tumor model. The enhanced stability provided by doubly strapped molecular design will enable new classes of in vivo NIR fluorescence imaging experiments with possible translation to humans.

Water-soluble organic macrocycles based on dye chromophores and their applications

“…21 Soon after, two doubly strapped heptamethine cyanine dyes were developed as NIR fluorescence labels, showing much enhanced stability. 46 Thus, this strategy provides a promising avenue to water-soluble dye-insulated macrocycles for biomedical applications, and it is expected that more double-cavity macrocycles with different dye cores will be exploited.…”

“…9b). 46 Compared to the parent dye, the doubly strapped 17 and 18 exhibit substantially greater chemical and photo-stability, which is attributed to the steric shielding from a macrocyclic structure. More interestingly, 17 and 18 with an unbound carboxyl group can be conjugated to antibodies, and the fluorescent antibody conjugates show excellent tumor-targeting specificity in living mice.…”

Engineered aptamers for molecular imaging

“…To overcome the aforementioned limitations, one strategy involves the utilization of near-infrared (NIR) fluorophores with longer excitation and emission wavelengths, enabling enhanced tissue penetration and reduced autofluorescence background. 157–160 For example, Hu et al developed and employed an integrated visible and NIR-I/II multispectral imaging system, which demonstrated enhanced contrast, sensitivity, and accuracy for the fluorescence imaging of deep tissues using the second NIR window (NIR-II, 1000–1700 nm). 161 Sicco et al utilized Alexa647-modified Sgc8c aptamers with far-red fluorescence to investigate the uptake characteristics of the probe in a murine metastatic melanoma model.…”

Design of Water–Soluble Rotaxane-Capped Superparamagnetic, Ultrasmall Fe3O4 Nanoparticles for Targeted NIR Fluorescence Imaging in Combination with Magnetic Resonance Imaging

“…This urges researchers to keep developing more innovative and comprehensive imaging techniques. In comparison to UV and visible light, near-infrared (NIR) light excitation (650–900 nm) has certain benefits in the form of deep tissue penetration, little background noise, and negligible autofluorescence signal from intracellular biomolecules that proffers an impressive tool for high-performance cellular organelle imaging. , Moreover, NIR fluorescence (NIR-FL) exhibits high sensitivity at the molecular level with fast feedback along with temporal/spatial resolution . However, NIR-FL imaging also has its inherent limitation, such as it is unable to provide anatomical 3D structural information.…”