Aminoglycosides restore full-length type VII collagen by overcoming premature termination codons: therapeutic implications for dystrophic epidermolysis bullosa.

[dystrophic epidermolysis bullosa]

Patients with recessive dystrophic epidermolysis bullosa (RDEB ) have severe , incurable skin fragility , blistering , and multiple skin wounds due to mutations in the gene encoding type VII collagen (C 7 ), the major component of anchoring fibrils mediating epidermal-dermal adherence . Nearly 10 -25 % of RDEB patients carry nonsense mutations leading to premature stop codons (PTCs ) that result in truncated C 7 . In this study , we evaluated the feasibility of using aminoglycosides to suppress PTCs and induce C 7 expression in two RDEB keratinocyte cell lines (Q 251 X /Q 251 X and R 578 X /R 906 ) and two primary RDEB fibroblasts (R 578 X /R 578 X and R 163 X /R 1683 X ). Incubation of these cells with aminoglycosides (geneticin , gentamicin , and paromomycin ) resulted in the synthesis and secretion of a full-length C 7 in a dose-dependent and sustained manner . Importantly , aminoglycoside-induced C 7 reversed the abnormal RDEB cell phenotype and incorporated into the dermal-epidermal junction of skin equivalents . We further demonstrated the general utility of aminoglycoside-mediated readthrough in 293 cells transiently transfected with expression vectors encoding 22 different RDEB nonsense mutations . This is the first study demonstrating that aminoglycosides can induce PTC readthrough and restore functional C 7 in RDEB caused by nonsense mutations . Therefore , aminoglycosides may have therapeutic potential for RDEB patients and other inherited skin diseases caused by nonsense mutations .