Engineering protein nanocages as carriers for biomedical applications

Bhaskar, Sathyamoorthy; Lim, Sierin

toimmune diseases. 8 Other ligands used for biological targeting include antibodies, engineered antibodies (e.g. single-chain variable fragment) and non-natural peptide ligands. Non-natural peptide ligands are often identified using phage-display libraries that provide the opportunity to screen multiple ligands with different affinities towards a specific cell receptor. Aptamers are nucleotide-based molecules that have been garnering interest as targeting ligands. The targeting ability of a carrier depends mainly on its surface ligand density, which can be tuned on protein nanocages to suit a given application. 8, 45 ENGINEERING OF PROTEIN SCAFFOLDS FOR SPECIFIC BIOMEDICAL APPLICATIONS Similar to other nanoparticles, the application of protein nanocages in biomedicine involves several challenges: (1) lack of a natural capability to carry drugs, (2) lack of specificity, (3) low cell uptake efficiency, (4) absence of endosomal escape mechanism, (5) limited circulation time, (6) potential to trigger an immunological response and (7) lack of tuneable release properties. Furthermore, clinical applications of nanocages are limited because of the structures' poor stability and cell permeability. 2 To overcome some of these challenges, the engineering of protein nanocages is required to impart non-natural functions. For example, the translocation of these carriers into cells through endocytosis is not always advantageous as these materials are digested in the lysosome rather than shuttled into the target cell organelle. Hence, incorporating an endosomal escape mechanism into protein nanocages is beneficial. What distinguishes nanocage-based delivery systems from other inorganic nanoparticles is the spatial control of functional groups and the ligands attached to the protein structure. Multifunctional properties can be achieved by combining the desired modifications for loading, targeting and chimeric assembly, leading to the development of smart nanocarriers with tremendous potential in nanomedicine. 4 Selective covalent chemical modifications in protein nanocages are made by leveraging native amino acids such as lysine, glutamic acid, aspartic acid and cysteine. 7 Table 1 provides a library of natural protein cage structures that have been chemically or genetically engineered to impart functional groups to different cage surfaces, such as the interior, exterior or intersubunit interface.