Assessing Cellular Internalization Pathways for Lipid Coated Silica Nanoparticle Mediated Nucleic Acid Delivery Platforms

The use of nanotechnologies for delivery of nucleic acids in gene therapy is a promising approach for the treatment of diseases resulting from genetic mutations. While promising, this technology has many biological barriers for effective delivery of nucleic acids. More specifically, internalization and intracellular pathways in a cell dictate the ultimate fate of the nanoparticle and nucleic acids which impacts overall effectiveness of the therapeutic approach. Endocytosis is a path by which cells take in substances from outside and submerge them in a vesicle. Upon internalization on the plasma membrane, lipid-coated silica nanoparticles serving as nucleic acids carriers are intended to use this pathway to reach the cytosol and release the nucleic acid in the nucleus to target a specific gene of a disease. Silica is considered safe by the FDA and engineered lipid coatings are expected to improve the biocompatibility of the particles. Bare lipid-coated silica nanoparticles, however, often get trapped and then degraded by the enzymes contained in the endosomes/lysosomes, thus preventing the desired RNA from reaching its intended destination, meaning a modulation of the particles is needed to enable an endosomal escape. In our study, we investigated the internalization pathway of lipid-coated silica nanoparticles in LNCaP cells, a prostate cancer cell line. Our approach involves assessing pathways for various lipid formulations using flow cytometry, then modulating lipid coated silica nanoparticles to improve their characteristics using two different systems and look at the colocalization of those particles with endosomal markers using confocal microscopy to assess whether an endosomal escape occurs and how our lipid-coated silica nanoparticles are internalized.