A new paper published in the journal EMBO Molecular Medicine by a team from the Lausanne University Hospital describes a protocol that can ensure the safety of adult epidermal stem cells before they are used as treatments for patients. The approach devised by this team takes cultivated, genetically modified stem cells and isolates single cells that are then used to make clonal cell cultures. These cloned cells are then rigorously tested to ensure that they meet the highest possible safety criteria. This protocol was inspired by approaches designed in the biotechnology industry and honed by regulatory authorities for medicinal proteins produced from genetically engineered mammalian cells.
“Until now there has not been a systematic way to ensure that adult epidermal stem cells meet all the necessary requirements for safety before use as treatments for disease,” says EMBO Member Yann Barrandon, Professor at Lausanne University Hospital, the Swiss Federal Institute of Technology in Lausanne and the lead author of the study. “We have devised a single cell strategy that is sufficiently scalable to assess the viability and safety of adult epidermal stem cells using an array of cell and molecular assays before the cells are used directly for the treatment of patients. We have used this strategy in a proof-of-concept study that involves treatment of a patient suffering from recessive dystrophic epidermolysis bullosa, a hereditary condition defined by the absence of type VII collagen which leads to severe blistering of the skin.”
Barrandon and co-workers have cultivated epidermal cells from patients who suffer from epidermolysis bullosa. These cells were then genetically engineered in order to insert a normal copy of the type VIII collagen gene. Then the genetically fixed cells were grown in culture so that they can be used to regenerate skin. Barrandon and others subjected these cells to an array of tests in order to determine which of the genetically engineered cells meet the requirements for safety and “stemness,” which refers to the stem cell characteristics that distinguish it from regular cells; its developmental immaturity and its ability to grow and self-renew. Clonal analysis revealed that the cultured, genetically engineered stem cells varied in their ability to produce functional type VII collagen. When the most viable, modified stem cells were selected and transplanted into the skin of immunodeficient mice, the cells regenerated skin and produced skin that did not blister in the mouse model system for recessive dystrophic epidermolysis bullosa. Furthermore, the cells produced functional type VII collagen. The safety of the cells was assessed by mapping the sites of integration of the viral vector. Because such viruses and produce gene rearrangements other mutations, the chosen cell lines were subjected to whole genome sequencing. Only the cells with insertions in benign locations were considered for use in their mouse model.
Barrandon concluded: “Our work shows that at least for adult epidermal stem cells it is possible to use a clonal strategy to deliver a level of safety that cannot be obtained by other gene therapy approaches. A clonal strategy should make it possible to integrate some of the more recent technologies for targeted genome editing that offer more precise ways to change genes in ways that may further benefit the treatment of disease. Further work is in progress in this direction.”
This work is certainly fascinating, but I think that using integrating viral vectors is asking for trouble. Certainly it should be possible to fix or replace the abnormal type VII collagen gene. Viruses that randomly insert genes into the genome can cause genetic problems, and even sequencing the genome may not properly address the safety concerns of the use of such viral vectors.