In vivo delivery of antioxidant enzymes with multi-functionalized lipid nanoparticles for sepsis therapy

Supplementing the cell with specific proteins is essential for disease prevention and therapy. However, protein permeability to the cell membrane is quite low because of the molecules large size and hydrophilic nature. Although protein delivery systems have been developing using vectors, their protein encapsulation efficiency depends on electrostatic interaction between proteins and vectors. Since proteins have a weaker net charge high affinity between vector and protein cannot be realized, and thus the encapsulation efficiency of naked proteins into vector is low. Herein, we developed a strategy for delivering target proteins into cells utilizing multifunctionalized lipid nanoparticles (MF-LNPs) prepared using several functional lipids that induce noncovalent interactions. We used two types of antioxidant enzymes, superoxide dismutase (SOD) and catalase (CAT), as model proteins for inflammation therapy. MF-LNPs are prepared by combining positively charged, neutral, and hydrophobic group-functionalized lipids. Optimization of the functional lipid composition alone resulted in MF-LNPs exhibiting nM affinity for SOD and CAT. Proteins were encapsulated in each optimized MF-LNP by freeze-thawing of MF-LNP and protein complexes. Co-treatment with SOD- and CAT-encapsulated MF-LNPs significantly inhibited ROS production in vitro and in vivo. Notably, the survival rate of model mice with severe sepsis was dramatically improved following the co-administration of SOD- and CAT-encapsulated MF-LNPs through the suppression of inflammatory cytokines and improvement of ROS scavenging activities. The findings indicate that this fundamental technology could be useful information for designing protein delivery vectors.