FC‐27 A spontaneous dog model for in vivo gene therapy of junctional epidermolysis bullosa
Junctional epidermolysis bullosa (JEB) is a mechanobullous skin disease associated with mutations in the basement membrane components of the dermo-epidermal junction. This condition is a suitable prototype for proving the feasibility of a gene therapy approach to genodermatoses. As successful genetic treatment requires immunocompetent animal models to study the possible host immune response to the transgene, we have characterized a breed of short-haired pointers suffering from a mild form of JEB. These animals exhibit skin blistering and erosions following minor trauma. Immunohistochemical analysis of the skin biopsies and immunoprecipitation of spent medium from cultured JEB keratinocytes showed reduced expression and secretion of the α3 chain of laminin 5 (α3, β3, γ2), the major adhesion ligand of basal keratinocytes. The search for genetic mutations detected a homozygous insertional mutation (4818 + 207 ins6.5 kb) in intron 35 of the lama3 gene. Consequently, keratinocytes from dogs with JEB secrete reduced amounts of wild-type laminin 5, and adhesive properties of the keratinocytes are compromised. Retroviral transfer of wild-type dog α3 cDNA into JEB keratinocytes enhanced secretion of laminin 5 in the extracellular matrix and restored the adhesion, differentiation and proliferative capacity of the transduced cells. Transplantable fibrin-based skin equivalents made with transduced JEB keratinocytes and grafted onto SCID mice generated normal cohesive and stratified epithelia and showed stable localized deposition of laminin 5 at the dermo-epidermal junction. Our results form the basis for preclinical assays of gene therapy on a unique immunocompetent animal model for an inherited skin disease. Funding: DEBRA (UK) Foundation, Association Francaise Contre les Myopathies.