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An Engineered Lipoproteoplex Presents Robust Delivery Mechanism For Topical Gene Therapy
Piul S. Rabbani, PhD1, Joseph A. Frezzo, MS2, Maria Ham, MD1, April Duckworth, MD1, Muhammad Hyder Junejo, BS1, Nakul Talathi, None1, Camilo Doig-Acuna, BS1, Haresh More, MTech2, Kevin Zhang, None2, Jessica Chang, BS1, Karan Mehta, BS1, Amanda Hua, BS1, Jin K. Montclare, PhD2, Pierre B. Saadeh, MD1, Daniel J. Ceradini, MD1.
1New York University Department of Plastic Surgery, New York, NY, USA, 2Polytechnic Institute of New York University, New York, NY, USA.
Purpose: Dermal penetration, drug degradation/toxicity, and non-specific effects are challenges in implementation of topical drug delivery systems. Here, we examine a new topical siRNA delivery system using a self-assembly protein-lipid system (lipoproteoplex) comprised of cationic lipid nanoparticles (CLNs) and Cartilage Oligomeric Matrix Protein coiled coil supercharged protein (CSP) for in vivo gene silencing. CSP carries a cavity for encapsulation and release of many different therapeutic agents, as well as the ability to complex siRNA. We predict that CLN/CSP will allow efficient siRNA delivery into skin, as well as sustained presence for maximum therapeutic effect.
Methods: Using various combinations of CSP and CLN with siRNA, we determined the optimal zeta potential of the complex in comparison to commonly used liposomal delivery systems. To analyze delivery efficiency in vivo, we complexed CSP/CLN with fluorescent siGlo Red siRNA and applied topically to wild type mouse skin. The mice were imaged in real time at 0, 3, 5, and 7 days after application with the in vivo imaging system (IVIS) and their skin sampled for histology and gene expression. In vitro, we used a Franz diffusion cell to determine release kinetics of the CSP/CLN lipoproteoplex.
Results: Surface charge of CSP:CLN:siRNA measured 37.33 ± 0.67 mV compared to 25.4 ± 1.15 mV of CLN:siRNA lipoplex, indicating the lipoproteoplex is more stable and resists aggregation to have improved solubility. IVIS imaging of fluorescence from treated mouse skin revealed presence of siGlo Red beginning by day 3, and maintained at days 5 and 7 after application. Histologic examination demonstrated dermal presence of siGlo Red by day 3. We found presence of siGlo Red from epidermis to the panniculus carnosus by day 5 and the fluorescent signal was visible at 7 days after application in sections. Using several combinations of CSP and CLN, we found favorable release kinetics of the CSP:CLN:siRNA lipoproteoplex for topical use. There was no evidence of cutaneous inflammatory reactions grossly or in histologic analysis with either delivery system.
Conclusion: Our results demonstrate that the lipoproteoplex offers an efficient and reliable route to deliver siRNA for topical gene silencing. The stability and non-toxic nature of lipoproteoplex gives it a unique advantage and makes it an ideal candidate for clinical cutaneous drug delivery.
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