There is currently no approved treatment for Netherton syndrome, which is a severe, genetic skin disease characterized by chronic inflammation and excessive peeling of the skin. Even though the molecular disease mechanism has been assigned to the unopposed activity of the two tissue kallikrein-related peptidases 5 and 7 (KLK5 and KLK7) and has been validated in mouse models, no therapies specifically targeting these two proteases exist or are currently in clinical development. To address this, we have used phage display of cyclic peptides to develop highly potent and selective macrocyclic inhibitors of KLK5 (Ki = 1 nM) and KLK7 (Ki = 7 nM). To equip these inhibitors with suitable pharmacokinetic properties for systemic in vivo application, we applied a half-life extension strategy to the peptides. This improved the plasma stability of the inhibitors from a half life of less than 5 min to more than 90 hours in vitro and prolonged the elimination half-life to 7 hours in vivo. The inhibitors diffused surprisingly well into the skin and reached high concentrations in the epidermis. Efficacy studies in mouse models of Netherton syndrome are currently being performed by a collaborator and look promising. Thus, the molecules have the potential to be further development toward the first targeted therapy for Netherton syndrome.