Project Summary Netherton syndrome (NS) is a severe genetic disorder that affects the skin, hair, and immune system. NS is caused by mutations associated with the SPINK5 gene which encodes LEKTI (Lympho-epithelial Kazal-type inhibitor), a serine protease inhibitor. Normally, skin shedding is controlled by an extracellular balance between serine proteases and serine protease inhibitors, and mutations in LEKTI result in overactivity of serine proteases. There is not an FDA-approved therapy or consensus treatment of conditions, and symptoms are mostly managed through antibacterials, emollients and corticosteroids. An enzyme-replacement therapy of topically applied LEKTI could be an elegant solution, but the short half-life of LEKTI precludes this possibility. The skin microbiome is home to over 1011 bacteria, and provides an attractive target as a delivery vehicle through continuous delivery of LEKTI. Through synthetic biology, we will engineer a bacteria to colonize the skin surface and secrete bioactive LEKTI to restore balance to the skin shedding process. In this Phase I SBIR, we will establish the safety of our topical application in in vitro models and demonstrate bioactive secretion of LEKTI fragments. This will provide a strong foundation to transition to Phase 2 where we will establish safety and efficacy in murine genetic models.