594 Gentamicin therapy induces functional laminin 332 and improves wound healing in junctional epidermolysis bullosa patients harboring nonsense mutations

Abstract

Herlitz Junctional Epidermolysis Bullosa (H-JEB) is an incurable and fatal inherited blistering skin disease most commonly caused by nonsense mutations in the LAMA3, LAMB3, or LAMC2 genes. These mutations impair production of functional laminin 332, which is needed for epidermal-dermal adherence. Previously, we demonstrated in vitro that gentamicin induced readthrough of premature stop codons (PTC) in H-JEB cultured keratinocytes harboring nonsense mutations and restored functional laminin 332. Herein, we examined if topical gentamicin could induce PTC readthrough and create new laminin 332 in H-JEB patients. H-JEB patients with nonsense mutations in LAMA3 and LAMB3 applied 0.5% gentamicin ointment topically to open wound Test sites daily for 14 days. Patient skin biopsies were examined before and at one and three months after treatment for the expression of laminin 332 and α6β4 integrin. Wounds were evaluated using photographs taken at zero, one and three months after treatment and Test Sites were assessed for new blister formation based on patient diaries and weekly questionnaires. Gentamicin treated sites exhibited newly created, correctly localized laminin 332 at the dermal-epidermal junction of their skin. The restoration of laminin 332 at these sites varied between 40% and 60% of the laminin 332 expression found in normal human skin and was durable for at least three months. Moreover, restored laminin 332 resulted in significantly increased expression and polarization of β4 integrin (from baseline 30% to 100% at 1 month and 3 months). Topical gentamicin corrected dermal-epidermal separation, improved wound closure and reduced new blister formation. No adverse events were observed. In summary, topical gentamicin reads through nonsense mutations in H-JEB patients and creates new laminin 332. Gentamicin therapy may provide a novel, low cost, non-invasive, and readily available therapy for JEB patients with nonsense mutations.