Ratiometric fluorescence monitoring of antibody-guided, targeted drug delivery to cancer.

Targeted drug delivery (TDD) systems, consisting of a drug or prodrug bound to a target-specific carrier such as an antibody, peptide, or nanoparticle, facilitate drug delivery, minimize side-effects to normal cells and organs, and thereby help to improve the safety of treatment including cancer. TDD systems additionally equipped with a switchable fluorescent dye bound to the drug or prodrug through a cleavable linker provide real-time monitoring of drug distribution and accumulation, as well as detection and diagnosis of abnormal cells. Upon the environment-mediated cleavage of the linker, the non-fluorescent switchable dye becomes fluorescent, signaling the drug release. The fluorescence signal from the dye is, however, affected by the light absorption and scattering in the biological sample, in particular, in the body, and is also dependent on the instrumental setup. Ratiometric measurements utilizing two independent fluorescence signals from a dual-dye conjugates are known to improve the detection sensitivity and quantification in many analytical, bioanalytical and pharmaceutical assays, including diagnostics and drug delivery monitoring. Nevertheless, these dual-dye conjugates have never been utilized for ratiometric monitoring of antibody-guided targeted drug delivery.

Here we report on the new, dual-dye TDD system Aza-Flu-Ab-Cy5s, comprising the switchable fluorescein-based dye (Flu) linked to the anticancer drug azatoxin (Aza), reference heptamethine cyanine dye (Cy5s), and Her2-specific humanized monoclonal Trastuzumab (Herceptin) antibody (Ab). A ratiometric fluorescence monitoring of drug release was demonstrated with this system in the example of the human breast cancer SKBR3 cell line overexpressing Her2 receptors.

Importantly, the developed ratiometric TDD system can further be modified for diagnosis and treatment of other types of cancer, which can be done by replacing the antibody and the drug molecules. The switchable and reference dyes can also be replaced to adjust the detection spectral range.