Targeted cellular delivery of robust enzyme nanoparticles for the treatment of drug-induced hepatotoxicity and liver injury.

Direct delivery of proteins into cells has been considered an effective approach for treating the protein-related diseases. However, clinical use of proteins has still been limited due to their instability in the blood and poor membrane permeability. To achieve an efficient cellular delivery of the protein to target cells via a systemic administration, a multifunctional carrier system having desirable stability both in the blood stream and the cells, specific cell-targeting property and endosomal escape functions may be required. In this study, we prepared a catalytic nanoparticle containing an active enzyme by cross-tethering multiple superoxide dismutase (SOD) molecules with catechol-derivatized hyaluronic acid (HA). The permeable shell of hydrophilic HA chains effectively protects the enzyme from degradation in the blood after intravenous administration and provides an additional function for targeting hepatocytes expressing HA receptor (CD44). The structure and catalytic activity of the enzyme molecules in the nanoparticle were not significantly compromised in the nanoparticle. In addition, ultra-small calcium phosphate nanoparticles (USCaP, 2-5 nm) were crystalized and decorated on the surface of the nanoparticle for the efficient endosomal escape after cellular uptake. The SOD-containing nanoparticle fortified with USCaP was used for the treatment of acetaminophen (APAP)-induced fulminant hepatotoxicity and liver injury. The nanoparticle achieved the efficient hepatic cellular delivery of SOD via a systemic administration and resulted in efficient removal of reactive oxygen species (ROS) in the liver and remarkable improvement of APAP-induced hepatotoxicity and liver injury in animals. STATEMENT OF SIGNIFICANCE: Despite the enormous therapeutic potential, the intracellular delivery of proteins has been limited due to their poor membrane permeability and stability. In this study, we demonstrated an active enzyme-containing nanoparticle functionalized by hyaluronic acid and ultra-small size calcium phosphate nanoparticles (2-5 nm) for targeted cellular delivery of superoxide dismutase (SOD). The nanoparticle was designed to integrate all the essential functions, including serum stability, target specificity, and endosomal escape capability, for a systemic delivery of a therapeutic protein to the cells of the liver tissue. The intravenous administration of the nanoparticle efficiently removes reactive oxygen species (ROS) in the liver and remarkably improves the drug-induced hepatotoxicity and the progress of fulminant liver injury in an acetaminophen-overdose animal model.