TARGET EDITING OF THE WISKOTT-ALDRICH SYNDROME (WAS) GENE USING ZINC FINGER NUCLEASES

Abstract

Wiskott-Aldrich syndrome (WAS) is an X-linked recessive disorder characterized by thrombocytopenia with small-sized platelets, recurrent infections, eczema, and increased susceptibility to autoimmune diseases and hematologic malignancies. It is caused by mutations in the gene encoding the WAS protein (WASP), a regulator of actin cytoskeleton and chromatin structure. The WASP is expressed in all hematopoietic lineages. Patients with severe WAS mostly die from infection or bleeding within the first decades of life. Hematopoietic stem cell transplantation (HSCT) remains the only curative therapy for WAS. The difficulty in finding HLA-matched donors and complications related to transplantation urge the development of gene therapy strategies for WAS. With the improvement in techniques for generating clinical-grade hematopoietic stem cells from induced pluripotent stem cells (iPSCs) and advances in gene targeting strategies, gene-corrected hematopoietic stem/progenitor cells derived from the patient iPSCs could be an effective alternative for treatment of severe immunodeficiency disorders. Here we reported on the successful generation of the genetically-corrected patient-specific WAS-iPSCs using zinc finger nuclease-based strategies. A total of eight corrected WAS-iPSC clones were obtained and revealed no random integration. These corrected WAS-iPSCs could be further validated prior to use in a clinical setting. This study is one of the most important steps in using gene targeting strategies to precisely correct the WASP mutation in patient-derived iPSCs.