Multimodal functional nanoparticles for applications in Nanotherapeutics

In this Work, the synthesis and preliminary optimization of a multimodal Au@SiO2 nanoparticle system for correlative studies into bio nano interactions and in vitro/in vivo biodistribution were developed. Multiple reporter modalities were built into the nanoparticle design namely two different Cyanine dyes that would allow for fluorescence cell and live animal imaging. The gold core would also provide easy tracking of the nanoparticles in vivo via mass spec methods which could be correlated at the whole organism and tissue level with the fluorescence. The gold core also acted as a contrast agent in electron microscopy of cell samples which was correlated with fluorescence by using CLEM microscopy workflows. These multimodal Au@SiO2 nanoparticles. The surface of these nanoparticles was functionalized with different Polyethylene glycol linkers via amination of the silica shell surface. Careful attention was given to developing methods for the surface functionalization of these nanoparticles which minimized any aggregation of the nanoparticles. The resulting PEGylated nanoparticles facilitated the bioconjugation of an Apo-E-CT receptor binding domain fragment to their surface in a range of ligand densities. This provided the opportunity to probe two questions, 1) role of orientation of the fragments on the surface effect on receptor recognition and 2) role of ligand density on recognition. A library of Apo-E-CT Au@SiO2 bionanoconstructs were developed which were studied using a QCM bionanoconstruct-receptor binding assays. These studies provided an insight into how these bionanoconstructs were recognized by three model receptors (Apo-E R2, LDLR and MARCO) ex vivo. These studies highlighted the role of orientation and the importance of building in studies of this nature in parallel with immuno gold mapping methods for availability of recognition motif is necessary to developing future nanomaterials for targeted nanodevices. The in vitro and in vivo evaluation of the non-functionalized Au@SiO2 nanoparticles was conducted to evaluate their performance in studying nanoparticle biodistribution and would serve to benchmark future studies into the biodistribution of the Apo-E bionanoconstructs. This work demonstrates the advantages of using a multimodal nanoparticle system for the study of ligand-receptor interactions, intracellular trafficking and Biodistribution. These materials are powerful tools for the correlation of findings across biological experiments from ex vivo ligand recognition studies to in vitro intracellular trafficking studies and then in vivo animal model biodistribution studies. They provide a strong platform in conjunction with established characterization workflows outlined throughout this thesis to allow for ranging comprehensive investigations into a deeper understanding and synthesis of potential novel nanotherapeutics.