Existing transdermal delivery systems (TDDS) for genetic diseases remain limited by poor transient gene expression and restricted patient accessibility. Microneedle patches, while minimally invasive, are hindered by complex mechanical fabrication, low cargo capacity, and limited shelf-life stability. Viral vectors, though effective, are costly to manufacture, have low copy-number capacity, and pose risks for adverse immune reactions. In 2023, the FDA approved B-VEC, an HSV-1–based vector delivering COL7A1 for Dystrophic Epidermolysis Bullosa (DEB), marking a milestone in topical gene therapy. However, its ~$600,000 annual cost underscores the urgent needs for scalable, shelf-stable, and cost-effective TDDS for long-term use and broader global accessibility. We simultaneously address transdermal and intercellular delivery using a localized injection of poly(beta-amino-ester) (PBAE) nanoparticles, formed through the electrostatically induced self-complexation of highly cationic PBAE polymers with a nucleic acid cargo. PBAEs carry 10-100X the plasmid copy number as viral vectors, are biodegradable, and can be synthesized with cheap commercially available reagents. We have developed a large library of PBAEs spanning different architectures (linear and branched) and chemical properties (hydrophilic and lipophilic), that when complexed with reporter GFP and delivered in vitro, have shown successful transfection (>60%) in hDFb fibroblasts and HacaT keratinocytes. A select few highest-performing linear and branched polymers were chosen for translation in vivo with a reporter fLuc plasmid, and gene delivery and expression were verified with IHC-HRP staining of FFPE skin sections. Encouraged by strong in vivo signal, we next focused on improving nanoparticle stability and storage through lyophilization. Due to the harsh nature of freeze-drying, we found that a 10% sucrose concentration acts as a necessary cryoprotectant. Additionally, we systematically optimized buffer ion valency and pH, nanoparticle concentration, and final resuspension buffer, and looked at stability via Dynamic Light Scattering (size, zeta potential, and PDI) post-lyophilization. Future work will include further optimizing lyophilization conditions to enhance in vitro to in vivo translation and to create a long-term shelf-stable nanoparticle, as well as combining PBAEs with a less invasive transdermal delivery method, such as STAR particles or microneedles to make micro-abrasions in the skin, for direct delivery of nanoparticle to the intercellular space of interest.
