Chamorro, CristinaMencía Rodríguez, ÁngelesAlmarza, DavidDuarte, BlancaBüning, HildegardSallach, JessicaHausser, IngridRío Nechaevsky, Marcela delLarcher Laguzzi, FernandoMurillas, Rodolfo2023-11-212023-11-212016-01Chamorro, C., Mencía, Á., Almarza, D., Duarte, B., Büning, H., Sallach, J., Hausser, I., Del Río, M., Larcher, F., & Murillas, R. (2016). Gene editing for the efficient correction of a recurrent COL7A1 mutation in recessive dystrophic epidermolysis bullosa keratinocytes. Molecular Therapy - Nucleic Acids, 5, e307.2162-2531https://hdl.handle.net/10016/38921Clonal gene therapy protocols based on the precise manipulation of epidermal stem cells require highly efficient gene-editing molecular tools. We have combined adeno-associated virus (AAV)-mediated delivery of donor template DNA with transcription activator-like nucleases (TALE) expressed by adenoviral vectors to address the correction of the c.6527insC mutation in the COL7A1 gene, causing recessive dystrophic epidermolysis bullosa in a high percentage of Spanish patients. After transduction with these viral vectors, high frequencies of homology-directed repair were found in clones of keratinocytes derived from a recessive dystrophic epidermolysis bullosa (RDEB) patient homozygous for the c.6527insC mutation. Gene-edited clones recovered the expression of the COL7A1 transcript and collagen VII protein at physiological levels. In addition, treatment of patient keratinocytes with TALE nucleases in the absence of a donor template DNA resulted in nonhomologous end joining (NHEJ)-mediated indel generation in the vicinity of the c.6527insC mutation site in a large proportion of keratinocyte clones. A subset of these indels restored the reading frame of COL7A1 and resulted in abundant, supraphysiological expression levels of mutant or truncated collagen VII protein. Keratinocyte clones corrected both by homology-directed repair (HDR) or NHEJ were used to regenerate skin displaying collagen VII in the dermo-epidermal junction.13eng© 2016 Official journal of the American Society of Gene & Cell Therapy. Published by Elsevier Inc.Atribución-NoComercial-CompartirIgual 3.0 EspañaAAV vectorsEpidermolysis bullosaGene editingIndelsHomologous recombinationTalenGene editing for the efficient correction of a recurrent COL7A1 mutation in recessive dystrophic epidermolysis bullosa keratinocytesresearch articleBiología y BiomedicinaInformáticaMedicinahttps://doi.org/10.1038/mtna.2016.19open access1e30713Molecular Therapy-Nucleic Acids5AR/0000019340