A single epidermal stem cell strategy for safe ex vivo gene therapy by Stéphanie Droz ‐ Georget Lathion, Ariane Rochat, Graham Knott, Alessandra Recchia, Danielle Martinet, Sara Benmohammed, Nicolas Grasset, Andrea Zaffalon, Nathalie Besuchet Schmutz, Emmanuelle Savioz ‐ Dayer, Jacques Samuel Beckmann, Jacques Rougemont, Fulvio Mavilio, and Yann Barrandon EMBO Mol Med. Volume ():e March 11, 2015 ©2015 by European Molecular Biology Organization

Strategy to perform ex vivo gene therapy from a single epidermal stem cellSchematic strategy to produce a performant and safe gene therapy product from a single autologous epidermal stem cell. Stéphanie Droz ‐ Georget Lathion et al. EMBO Mol Med. doi: /emmm ©2015 by European Molecular Biology Organization

Extensive growth potential of recessive dystrophic epidermolysis bullosa (RDEB) epidermal keratinocytes Keratinocytes were isolated from the skin of a 4 ‐ year ‐ old patient with severe ‐ generalised RDEB linked to homozygous insertion–deletion in COL7A1 (Hilal... Stéphanie Droz ‐ Georget Lathion et al. EMBO Mol Med. doi: /emmm ©2015 by European Molecular Biology Organization

Isolation of genetically corrected recessive dystrophic epidermolysis bullosa (RDEB) epidermal stem cellsSingle cells were isolated from a mass culture (passage V) of RDEB keratinocytes infected with SIN retroviruses bearing a COL7A1 cDNA. Stéphanie Droz ‐ Georget Lathion et al. EMBO Mol Med. doi: /emmm ©2015 by European Molecular Biology Organization

Long ‐ term restoration of COLVII expression, generation of epidermis and anchoring fibrils by the progeny of a corrected recessive dystrophic epidermolysis bullosa (RDEB) epidermal stem cell Serial cultivation of transduced and untransduced holoclones demon... Stéphanie Droz ‐ Georget Lathion et al. EMBO Mol Med. doi: /emmm ©2015 by European Molecular Biology Organization

Characterisation of proviral integrations in corrected stem cells Visualisation of COL7A1 sequence by FISH analysis in corrected stem cells at passage XII (clone 6). Stéphanie Droz ‐ Georget Lathion et al. EMBO Mol Med. doi: /emmm ©2015 by European Molecular Biology Organization

Assessment of immortalisation, tumorigenicity and disseminative potential of the corrected stem cells Upper panel: expression of proteins associated with the acquisition of immortalisation process involved in senescence, in evasion of growth suppression a... Stéphanie Droz ‐ Georget Lathion et al. EMBO Mol Med. doi: /emmm ©2015 by European Molecular Biology Organization