Year 1

The goal of this proposal is to complete the work necessary to file a pre-IND with the FDA focused on using genome editing of autologous blood stem cells to cure severe combined immunodeficiency caused by mutations in the IL2RG gene. In the first year of the project, we have significant progress. This progress includes:
1. Identifying that the CRISPR/Cas9 nuclease system is more efficient at editing the IL2RG gene than the TALENs we had previously made.
2. That combining recombinant AAV6 transduction and electroporation of the CRISPR/Cas9 ribonucleoprotein complex, we can achieve greater than 30% functional gene correction in blood stem and progenitor cells. Since experiments of nature demonstrate that single corrected progenitor cell can give rise to a functionally diverse immune system, the rate of editing that we are achieving exceeds our original goal.
3. That the functionally corrected blood stem and progenitor cells are able to give rise to the different blood types in the same amounts and the same proportions as un-manipulated cells.
4. That T-cells that have been modified using the functional gene correction approach are equivalent to wild-type T-cells in both their proliferative capacity and their ability to respond to cytokines.
5. That the CRISPR/Cas9 nuclease delivered into blood stem and progenitor cells is highly specific.

In sum, we made excellent progress and are in the process of performing the final animal transplantation experiments to determine if the functionally corrected blood stem and progenitor cells can reconstitute the blood system. If these transplant experiments are successful, which we believe they will be because of our results using the same system editing a different gene, then we will be in excellent position to begin putting together our pre-IND application for the FDA.