Endothelial delivery of antioxidant enzymes loaded into non-polymeric magnetic nanoparticles

Abstract

Antioxidant enzymes have shown promise as a therapy for pathological conditions involving increased production of reactive oxygen species (ROS). However the efficiency of their use for combating oxidative stress is dependent on the ability to achieve therapeutically adequate levels of active enzymes at the site of ROS-mediated injury. Thus, the implementation of antioxidant enzyme therapy requires a strategy enabling both guided delivery to the target site and effective protection of the protein in its active form. To address these requirements we developed magnetically responsive nanoparticles (MNP) formed by precipitation of calcium oleate in the presence of magnetite-based ferrofluid (controlled aggregation/precipitation) as a carrier for magnetically guided delivery of therapeutic proteins. We hypothesized that antioxidant enzymes, catalase and superoxide dismutase, can be protected from proteolytic inactivation by encapsulation in MNP. We also hypothesized that catalase-loaded MNP applied with a high-gradient magnetic field can rescue endothelial cells from hydrogen peroxide toxicity in culture. To test these hypotheses, a family of enzyme-loaded MNP formulations were prepared and characterized with respect to their magnetic properties, enzyme entrapment yields and protection capacity. SOD-and catalase-loaded MNP were formed with average sizes ranging from 300 to 400 nm, and a protein loading efficiency of 20-33%. MNP were strongly magnetically responsive (magnetic moment at saturation of 14.3 emu/g) in the absence of magnetic remanence, and exhibited a protracted release of their cargo protein in plasma. Catalase stably associated with MNP was protected from proteolysis and retained 20% of its initial enzymatic activity after 24 hr of exposure to pronase. Under magnetic guidance catalase-loaded MNP were rapidly taken up by cultured endothelial cells providing increased resistance to oxidative stress (62±12% cells rescued from hydrogen peroxide induced cell death vs. 10±4% under non-magnetic conditions). We conclude that non-polymeric MNP formed using the controlled aggregation/precipitation strategy are a promising carrier for targeted antioxidant enzyme therapy, and in combination with magnetic guidance can be applied to protect endothelial cells from oxidative stress mediated damage. This protective effect of

Restenosis Treatments Using Nanoparticle-based Drug Delivery Systems

“…This same approach was used to magnetically target antioxidant enzymes to endothelial cells to protect them from oxidative stress-mediated damage [104]. These results indicate that magnetic nanoparticles with magnetic guidance may be interesting candidates for stent-targeted anti-restenotic therapies.…”

“…Chorny and co-workers [102][103][104] demonstrated that magnetic field controlled targeting of paclitaxel-loaded magnetic nanoparticles (~250 nm in diameter)…”

Synthesis, Functionalization, and Design of Magnetic Nanoparticles for Theranostic Applications

“…[638] Moreover, MNPs have been employed for delivering antioxidant enzymes to increase resistance to oxidative stress. [639] In contrast to permanent magnets which take advantage of the guidance response taken by MNPs, an oscillating or alternating magnetic field provides a novel on-off fashion for triggering drug release. In fact, by employing thermo-responsive polymers [640] and lipids [641] onto the surface of MNPs and exploiting the hyperthermia effect the energy transformed into heat through Brownian fluctuations and Neel fluctuations can lead to promising magnetic responsive drug delivery systems.…”

Magnetic nanoparticles for targeted vascular delivery

“…Cellular uptake of MNP-Ad affinity complexes by cultured vascular cells and the resultant transduction efficiency was determined after cell treatment with or without a high-gradient The feasibility of a different formulation strategy developed for encapsulation of biotherapeutics in MNP under mild conditions without the use of organic solvents was recently demonstrated with the antioxidant enzymes, catalase and superoxide dismutase, and the protective efficiency of MNP-encapsulated catalase was shown in cultured endothelial cells challenged with hydrogen peroxide (56). This formulation method, termed controlled aggregation/precipitation, is potentially applicable to both proteins and gene vectors and is based on the coentrapment of a macromolecule and nanocrystalline magnetite in the matrix of MNP formed by precipitation of calcium or zinc salts of oleic acid in the presence of a surface stabilizer, such as Poloxamer 407.…”