Optimization of catalase-loaded liposomal magnesium phosphate nanoparticles for intracellular protein delivery
Poster Number
12
Introduction/Abstract
Protein therapeutics have great potential in treating human disease. Compared to small molecule drugs, proteins as therapeutic agents carry the potential advantages of exerting more complicated activities, higher specificity and higher biocompatibility. However, clinical use of intracellular proteins is hampered by their physicochemical properties such as large size, excess surface charge and relatively unstable structure. Such properties tend to induce their rapid denaturation and/or degradation, extensive uptake by the reticuloendothelial system (RES), and/or elimination by renal filtration. Furthermore, cell membranes also impose a formidable barrier for such proteins to cross in order to reach their intracellular target site. Apart from that, Proteins that enter cells through endocytosis will be eventually degraded in lysosomes if they do not escape the endosomal pathway before reaching lysosomes. Therefore, the development of protein delivery systems, including liposomal and/or polymeric nanoparticles would substantially facilitate the use of proteins in clinic.
Purpose
Optimization of cationic lipid-coated magnesium phosphate nanoparticle (LP MgP NP) formulations to reduce the nanoparticle size for efficient intracellular protein (catalase) delivery
Method
Magnesium phosphate nanoparticles (MgP NP) were prepared by precipitation of magnesium chloride and diammonium hydrogen phosphate micro-emulsions. Magnesium chloride and diammonium hydrogen phosphate micro-emulsions were sonicated individually using a probe sonicator (Branson digital sonifier), mixed together, and allowed to react. The resultant MgP NP in micro-emulsion was then sonicated and coated with sodium citrate to impart negative surface charge. The microemulsion was sonicated again and then mixed with an ethanol suspension of silica gel. The resultant silica gel coated with MgP NP was washed with ethanol and then eluted with ethanol/water mixtures. The eluents were evaporated to remove ethanol and yielded MgP NP in an aqueous suspension. Cationic liposomes of different compositions (DOTAP/cholesterol = 1:1, 6:4 and 7:3, mol/mol) were prepared by film hydration coupled with extrusion through membranes of different pore sizes. Cationic liposomes were complexed spontaneously with catalase upon mixing and the positively charged liposomecatalase complex was mixed with negatively charged MgP NP to yield the final formulation. Particle size was measured using Malvern NANO-ZS90 zeta sizer.
Results
Sonication at amplitude of 30 reduced the size of MgP NP to 345.1 nm (Image 1). Lipid films at 7:3 DOTAP/cholesterol molar ratio yielded cationic liposomes of 71.63 nm in diameter after extrusion through 50 nm membrane (Image 2), which then yielded liposome-catalase complex of 92.78 nm in diameter. After mixing the smallest MgP NP and the smallest liposome-catalase complex, the diameter of the optimized LP MgP NP formulation was reduced drastically to 102.3 nm (PDI = 0.173) from our prior preparations (D = 233.4 nm, PDI = 0.135).
Significance
Through sonication, extrusion and optimization of lipid composition, we have successfully prepared LP MgP NP formulations of reduced size which could facilitate efficient intracellular protein delivery.
Location
DeRosa University Center, Stockton campus, University of the Pacific
Format
Poster Presentation
Optimization of catalase-loaded liposomal magnesium phosphate nanoparticles for intracellular protein delivery
DeRosa University Center, Stockton campus, University of the Pacific
Protein therapeutics have great potential in treating human disease. Compared to small molecule drugs, proteins as therapeutic agents carry the potential advantages of exerting more complicated activities, higher specificity and higher biocompatibility. However, clinical use of intracellular proteins is hampered by their physicochemical properties such as large size, excess surface charge and relatively unstable structure. Such properties tend to induce their rapid denaturation and/or degradation, extensive uptake by the reticuloendothelial system (RES), and/or elimination by renal filtration. Furthermore, cell membranes also impose a formidable barrier for such proteins to cross in order to reach their intracellular target site. Apart from that, Proteins that enter cells through endocytosis will be eventually degraded in lysosomes if they do not escape the endosomal pathway before reaching lysosomes. Therefore, the development of protein delivery systems, including liposomal and/or polymeric nanoparticles would substantially facilitate the use of proteins in clinic.
