Engineering protein nanocages as carriers for biomedical applications

Bhaskar, Sathyamoorthy; Lim, Sierin

One of the key requirements of drug delivery systems is the effective loading and release of drugs from the nanoparticle carriers. The strategies used to choose a delivery system rely on the system's structure and functions, the nature of the drug carried and the microenvironment the drug encounters. The delivery of drugs/active molecules can be facilitated by chemical immobilization, non-covalent interaction and environment-dependent conformational changes of the carrier. Protein-based carriers and drugs could be chemically conjugated by post-translational attachment of drug molecules to reactive side chains of amino acids such as amines, carboxyls, sulfhydryls and hydroxyls, as well as non-side chains through click chemistry. Covalent conjugation allows for adequate control over release kinetics. The mode of drug release is chosen based on the conjugation chemistry between the drugs, the carrier and the cellular microenvironment encountered. The interior cavity of the protein nanocages often contains natural reactive sites for attachment of nucleic acids, drugs or metals. Drug molecules may also be loaded by nonspecific interactions with secondary carriers that have a strong affinity for the internal surface of the carrier. The nonspecific interactions may modulate drug release kinetics under physiological conditions. Diffusion through the native pores in the carrier facilitates entry into the central cavity. Gated pores in certain VLPs could swell open at low salt concentrations, high pH or osmotic shock, helping load the drug. When the conditions are reversed, the drug is retained, preventing outward diffusion. This mechanism can be applied to deliver drugs to acidic cancer microenvironments. The disassembly and reassembly of the protein cages based on environmental conditions could also aid drug loading and release. The release mechanism of loaded molecules could be tuned by introducing repulsive forces at the intersubunit interfaces to facilitate environment (e.g. pH)-triggered dissociation of protein subunits of the nanocarrier. 8, 43 Targeting Nanoparticles accumulate in the cancer microenvironment by enhanced permeation and retention (EPR) effect due to the leaky vasculature of the tissue. Nonspecific accumulation through the enhanced permeation and retention effect is referred to as passive targeting and may not be very effective. To enhance the affinity for target cells, the carriers can be decorated with targeting ligands to impart active targeting. Targeted delivery of drugs by carrier systems can reduce the amount of drug-carrier complex needed for therapy. An example of active targeting ligand is the peptide RGD, which is present in adenoviruses and has a natural affinity for upregulated integrin receptors in endothelial cells of tumor vessels. Attaching this peptide to the external surface of a carrier aids tumor-targeted delivery. Certain tumor delivery platforms focus on targeting the surface receptors using natural ligands that facilitate endocytosis, 44 Engineered protein cages in biomedical applications S Bhaskar and S Lim such as folate receptors in cancer cells, transferrin receptors and epidermal growth factor receptors. 40 Although there are many diseases that delivery systems target, research has focused on solid tumors, and cardiovascular diseases. One of the emerging areas of interest is the modulation of the immune response using protein carriers for application in tumor immunotherapy and the treatment of au