Gene Editing Technologies
The combination of gene-editing technologies alongside the use of stem cells for transplantation represents an attractive alternative to current approaches for disease treatment. Whereas a number of different approaches allowing for gene-targeting of stem cells have been described, the technologies available present important limitations. Random integration of DNA fragments poses a big concern. Besides their off-target effects, efficiency and carrying capacity is limited. Our goals involve the generation of novel technologies allowing not only for gene-correction but also for efficient gene-editing of multiple somatic cell types as well as pluripotent cells. Our interest include a broad range of strategies particularly focus to optimize error-free homologous recombination. As such our work currently covers the use of specific nucleases, such as Zinc Finger Nucleases (ZFN), as well as Bacterial Artificial Chromosomes (BAC) and the use of Helper Dependent Adenoviral vectors (HDAdV). Albeit each technology presents its cons and pros, their employment and further development could serve for a number of applications including introduction of suicide genes in safe-harbor genomic positions for the purging of residual pluripotent cells upon differentiation, the generation of reporter lines as well as the specific knock-in/out of selected genes for in-depth molecular studies. Eventually, our technologies could be expanded to a number of different fields and clinical studies relying on gene-editing such as targeted immunology and directed tumor-attack. In summary, we aim to address the following questions:
1) Are these technologies safe enough to guarantee clinical translation?
In order to translate gene-targeting methodologies to the clinic we aim to define a safety profile for each of the available or under development approaches. Factors such as the presence and number of random integrations have to be taken into account in order to avoid insertional mutagenesis leading to tumorigenesis. Moreover, even in the event of minimal number of integrated sequences into the host genome, the location of such exogenous sequences can have an impact on the actual safety profile of the modified cells. Additionally, we aim to investigate and define those technologies presenting the lowest toxicity as well as maintaining genomic and epigenomic integrity.
2) Can we optimize and/or develop approaches allowing for the correction of large genomic regions?
Even though variable, most of the current technologies present a DNA-carrying capacity which allows for the correction of single point mutations in the target gene. Yet, a number of different diseases are characterized by the presence of large duplications, translocations or deletions. We aim to expand the current approaches to allow for the correction of, not only single point mutations, but also larger genomic aberrations.
3) Is it possible to target adult stem cells?
A common limiting factor of most gene-editing technologies is the general inability for targeting adult stem cells. Yet, adult stem cells are a promising source of multi-potent progenitors with clinical implications, as evidenced by the increasing number of clinical trials involving the use of Mesenchymal Stem Cells. We are currently developing technologies directed to gene correction in adult stem cells, an approach that may facilitate patient autologous transplantation.