5'RNA -Splicing Repair of Mutant Transcripts in Epidermolysis Bullosa.

Mutations within the gene underlie the inherited recessive subtype of the blistering skin disease dystrophic epidermolysis bullosa (RDEB). Although gene replacement approaches for genodermatoses are clinically advanced, their implementation for RDEB is challenging and requires endogenous regulation of transgene expression. Thus, we are using spliceosome-mediated RNA -splicing (SMaRT) to repair mutations in at the mRNA level. Here, we demonstrate the capability of a -specific RNA -splicing molecule (RTM), initially selected using a fluorescence-based screening procedure, to accurately replace exons 1 to 64 in an endogenous setting. Retroviral RTM transduction into patient-derived, immortalized keratinocytes resulted in an increase in wild-type transcript and protein levels, respectively. Furthermore, we revealed accurate deposition of recovered type VII collagen protein within the basement membrane zone of expanded skin equivalents using immunofluorescence staining. In summary, we showed for the first time the potential of endogenous 5' -splicing to correct pathogenic mutations within the gene. Therefore, we consider 5' RNA -splicing a suitable tool to beneficially modulate the RDEB-phenotype, thus targeting an urgent need of this patient population.