Figure 3: Therapeutic CYBB Gene Correction.
X-CGD is caused by a mutation in the CYBB gene which encodes a critical component (gp91-phox) of NADPH oxidase, a key enzyme for the anti-microbial activity of phagocytes. Mutation correction offers a new curative treatment for X-CGD patients. The patient’s own cells are harvested, the mutated gene corrected using CRISPR-mediated gene editing, and the cells returned to the patient. The engrafted cells multiply to create a new population of cells displaying ‘normal’ function and eliminating disease. In these studies, CD34+ hematopoietic stem cells (HSPCs) were isolated from X-CGD patients and electroporated with CRISPR-Cas9, guide RNA and the gene-correcting, oligo template using the MaxCyte GTx. A portion of the cells were differentiated in vitro into myeloid cells and gene correction rates determined to be 31%. The other portion of corrected HSPCs were introduced into immunodeficient mice. After 20 weeks the engrafted human cells expressed the corrected gp91 gene at 14% in the mouse peripheral blood and 10% – 20% in the bone marrow. These correction rates are within clinically beneficial potency thresholds. Sci. Transl. Med., 9(372), Jan 2017.