Design of Protein-Derived Carbon Dots for Targeted Biomarker Labeling for Diagnostic Bio-Imaging and Targeted Therapeutics

Carbon dots (CDs) are a type of carbon-based nanoparticle possessing remarkable qualities such as unique optical properties, small size, high biocompatibility, and tunable surface chemistry. An important advantage of these nanoparticles is their ability to inherit the properties of the original carbon precursor. The retention of precursor chemistry enables directed design of the CDs final functionality, with no post synthesis modifications. This dissertation presents advancements in the largely unexplored research area of designing and implementing protein-derived CDs as a powerful analytical platform for bioimaging, biosensing, and targeted therapeutics. CD parameter optimization synthesis techniques were designed using bovine serum albumin as a model protein to enable rational design of protein-derived CDs with desired properties. This enables control of CDs photoluminescence emission, size, surface charge, and protein-ligand binding affinity. Fibronectin protein-derived CDs were developed as a histological stain for tumor lumpectomies modeled on HER2+ and triple negative breast cancer. The biocompatible NPs were brightly luminescent under UV light and selectively targeted fibronectin receptor, α5β1 integrin, which is often overexpressed in metastatic cancer cells. Due to CDs high quantum yield, tunable color emission, low cytotoxicity, incredibly small size, and intrinsic pre-designed affinity, protein-derived CDs are a powerful tool for super-resolution microscopy techniques (SRM) to elucidate altered biological mechanisms in malignant cells in vitro at the nanoscale. These particles can further be implemented to selectively interact with “undruggable” targets and induce changes in biological mechanisms. Transferrin-derived CDs were utilized to selectively bind cancer biomarker CD71 in promyelocytic leukemia cells. Super resolution radial fluctuation microscopy enabled nanoscale imaging of transferrin-derived CDs triggering CD71-mediated endocytosis. Particle tracking and mean square displacement calculations confirmed SRM images indicating that the CDs altered the cells recycling biomechanics resulting in an increased degradation pathway, retaining the NPs in the malignant cell’s lysosome. A CD71-affinity, drug delivering nanocarrier was designed with non-selective, apoptosis inducing drug, staurosporine, and transferrin-derived CDs. The resulting conjugate resulted in over 2X malignant cell apoptotic cell death. Protein-derived CDs are largely unexplored, however these advances in CD design have produced nanoparticles with advantages over current techniques including low cost, facile synthesis, and near limitless customizability via simple precursor selection. The most notable advantage is being able to attack previously considered “undruggable” intracellular targets for targeted therapeutics and theranostics.