Bone marrow or stem cell transplantation
CRISPR-based gene correction in hematopoietic stem and progenitor cells (HSPCs) has the potential to treat a wide variety of genetic and acquired diseases. Gene corrected HSPCs can be used for autologous transplantation, circumventing the need for HLA-matched donor transplants. Here we propose a CRISPR-based gene correction strategy to treat IPEX syndrome, the prototype of genetic autoimmunity. IPEX syndrome is caused by mutations in the FOXP3 gene, which leads to dysfunction of T regulatory cells (Tregs) and T effector (Teff) cells and subsequent autoimmune manifestations. Due to the widespread distribution of FOXP3 mutations throughout the gene, we designed a strategy to insert a FOXP3 cDNA into the mutated gene locus. Using the CRISPR system, we achieved efficient and specific targeting of FOXP3 in HSPCs, Tregs and Teff cells. Gene edited Teff cells maintained physiological regulation of FOXP3 expression and characteristic proliferation potential and cytokine production. Gene edited Tregs displayed partial FOXP3 protein expression and suppressive capacity relative to wild-type controls, and various cDNA constructs were compared in Tregs to optimize FOXP3 expression and function. Next, we showed that gene edited HSPCs retained the potential to differentiate in vitro and engraft and differentiate in immunodeficient mice. Lastly, we demonstrated that editing of IPEX cells is a feasible and efficient method for functional FOXP3 gene correction. This study supports a CRISPR-based strategy to treat IPEX syndrome, and further underscores the potential of gene editing treatments for other genetic immune diseases.