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One step closer to a CRISPR ex vivo gene therapy for DEB

New scientific publication from the Koller Working Group

Ex vivo gene therapy currently represents one of the most promising approach to permanently close chronic wounds, and thus preventing cancer formation in EB. A few junctional EB (JEB) patients with mutations in the laminin gene were successfully treated with a “conventional” ex vivo gene therapy, where patient’s own stem cells are corrected outside the body by using a virus to introduce a healthy copy of the mutated gene. The corrected stem cells are then grown into skin sheets for grafting onto wounds. Attempts to use the same protocol for recessive dystrophic EB (RDEB) patients did not result in long term wound closure. This might be due to the enormous size and special characteristics of the delivered collagen 7 gene, which impairs the delivery and insertion into stem cells and their efficient functioning after transplantation  and hence their long-term and stable correction.
Thus, for RDEB patients in particular, alternative strategies such as CRISPR-based gene editing represent a promising option for successful ex vivo gene therapy. CRISPR repairs the skin cells permanently by cutting out a small area of the mutated DNA using “gene scissors” and replacing it with a healthy copy, without the need for a virus. The method is very precise, but limited by the fact that only a small percentage of all treated cells are genetically corrected. For successful wound closure, the grafted skin must contain a high number of corrected skin stem cells. In the laboratory, CRISPR-corrected cells can be enriched (=selection) by inserting a gene for selection together with the repair molecule into the cell's genome. After adding a specific substance, only the corrected cells survive. However, the presence of a foreign gene in the cells is not feasible for a clinical application, since it could trigger undesirable immune reactions.

Researchers from the EB House have developed a CRISPR strategy that repairs mutations in EB cells in a specific manner and without the need for selection. In a current pre-clinical study, they have improved this approach in terms of efficiency and safety by exploiting novel technologies. Application of the new strategy in RDEB skin cells in the laboratory resulted in precise correction of the collagen 7 mutation in about 20 % of all treated cells, which exceeds all previous attempts. Successful repair was confirmed by the production of a healthy collagen 7 protein in cultivated skin cells and its deposition between the skin layers in a cultivated 3D skin. Although there are still some challenges to be overcome, the results give hope that, this ex vivo CRISPR gene therapy will be applied for DEB in the near future.

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