Protein cage architectures for targeted therapeutic and imaging agent delivery
Flenniken, Michelle Lynne.
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Protein cage architectures such as viral capsids, heat shock proteins, and ferritins are naturally occurring spherical structures that are potentially useful nanoscale platforms for biomedical applications. This dissertation work demonstrates the utility of protein cages including their use as therapeutic and imaging agent delivery systems. Protein cage architectures have clearly demarcated exterior, interior, and interface surfaces and their structures are known to atomic level resolution. This information is essential for the engineering of functionalized nanoparticles via both chemical and genetic modification. In the process of tailoring protein cage architectures for particular applications, fundamental information about the architectures themselves is gained. The present work describes endeavors toward the use of three different protein cage architectures, the Cowpea chlorotic mottle viral capsid (CCMV), a small heat shock protein (Hsp) architecture originally isolated from the hyperthermophilic archaeon Methanococcus jannaschii, and human H-chain ferritin, as cell-specific therapeutic and imaging agent delivery systems. Each protein cage is roughly spherical, but their sizes differ; CCMV is 28 nm in diameter, whereas Hsp and HFn are 12 nm in diameter.The advantages and disadvantages of all three architectures are described. Wild type and genetic variants of the Hsp, HFn, and CCMV cages were reacted for the site specific attachment of organic molecules such as therapeutic agents, imaging agents, and targeting ligands. Inorganic chemical modification of the cages was employed for the formation iron oxide nanoparticles which are potentially useful as magnetic resonance imaging (MRI) contrast agents. Toward the development of the Hsp platform for therapeutic delivery, the antitumor agent doxorubicin was covalently bound on the interior of the cage and selectively released via a pH dependant trigger. In addition, mammalian cell-specific targeting was imparted to the Hsp and HFn cages by both genetic and chemical strategies. Biodistribution studies of Hsp and CCMV were performed in naïve and pre-immunized mice and Hsp cages localized to human tumor xenografts in mice. Together these results demonstrate the utility of the protein cages as robust nanoscale platforms for the synthesis of both soft (organic) and hard (inorganic) materials.