Enhanced homology-directed repair for highly efficient gene editing in hematopoietic stem/progenitor cells
Suk See De Ravin, Julie Brault, Ronald J. Meis, Siyuan Liu, Linhong Li, Mara Pavel-Dinu, Cícera R. Lazzarotto, Taylor Liu, Sherry Koontz, Uimook Choi, Colin L. Sweeney, Narda Theobald, GaHyun Lee, Aaron B. Clark, Sandra Burkett, Benjamin P. Kleinstiver, Matthew H. Porteus, Shengdar Q. Tsai, Douglas B. Kuhns, Gary A. Dahl, Stephen J. Headey, Xiaolin Wu, Harry L. Malech
Abstract
Lentivector gene therapy for X-linked chronic granulomatous disease (X-CGD) has proven to be a viable approach, but random vector integration and subnormal protein production from exogenous promoters in transduced cells remain concerning for long-term safety and efficacy. A previous genome editing-based approach using Streptococcus pyogenes Cas9 mRNA and an oligodeoxynucleotide donor to repair genetic mutations showed the capability to restore physiological protein expression but lacked sufficient efficiency in quiescent CD34+ hematopoietic cells for clinical translation. Here, we report that transient inhibition of p53-binding protein 1 (53BP1) significantly increased (2.3-fold) long-term homology-directed repair to achieve highly efficient (80% gp91phox+ cells compared with healthy donor control subjects) long-term correction of X-CGD CD34+ cells.