PhiC31 integrase-mediated nonviral genetic correction of junctional epidermolysis bullosa.
Patients afflicted with severe laminin 5-deficient junctional epidermolysis bullosa (JEB) often die in infancy with massive cutaneous blistering. Prior approaches to genetically correct this disorder have relied on stable integration of wild-type LAMB3 sequences, using retroviral vectors. To develop a nonviral approach to JEB gene therapy, we used the phiC31 integrase, which mediates unidirectional genomic integration of plasmids containing a specific attB site. An attB-containing laminin 5 beta3 expression plasmid was integrated into the genomes of primary keratinocytes from four unrelated, genetically characterized JEB patients. phiC31 integrase supported genomic integration into epidermal progenitor cells. Regeneration of human skin on immunedeficient mice, using these cells, produced human skin tissue with restored laminin 5 expression. Furthermore, corrected JEB tissue restored hemidesmosome formation and abolished histologic evidence of subepidermal blistering. These findings provide an approach to durable nonviral correction of JEB.
Ortiz-Urda
Susana
S
VA Palo Alto Healthcare System and Program in Epithelial Biology, Stanford University School of Medicine, 269 Campus Drive, Stanford, CA 94305, USA.
Thyagarajan
Bhaskar
B
Keene
Douglas R
DR
Lin
Qun
Q
Calos
Michele P
MP
Khavari
Paul A
PA
D000818:Animals; D015815:Cell Adhesion Molecules / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D016109:Epidermolysis Bullosa, Junctional / Q000378:metabolism / Q000628:therapy*; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors; D006801:Humans; D019426:Integrases / Q000235:genetics*; D015603:Keratinocytes / Q000502:physiology / Q000648:ultrastructure; D051379:Mice; D016513:Mice, SCID; D010957:Plasmids; D012038:Regeneration; D012867:Skin / Q000033:anatomy & histology; D000097794:Kalinin
D015815:Cell Adhesion Molecules; D019426:Integrases; D000097794:Kalinin