TY - JOUR T1 - Efficient Correction of the Sickle Mutation in Human Hematopoietic Stem Cells Using a Cas9 Ribonucleoprotein Complex JF - bioRxiv DO - 10.1101/036236 SP - 036236 AU - Mark A. DeWitt AU - Wendy Magis AU - Nicolas L. Bray AU - Tianjiao Wang AU - Jennifer R. Berman AU - Fabrizia Urbinati AU - Denise P. Muñoz AU - Donald B. Kohn AU - Mark C. Walters AU - Dana Carroll AU - David K. Martin AU - Jacob E. Corn Y1 - 2016/01/01 UR - http://biorxiv.org/content/early/2016/01/15/036236.abstract N2 - Sickle Cell Disease (SCD) is a serious recessive genetic disorder caused by a single nucleotide polymorphism (SNP) in the ß-globin gene (HBB). Sickle hemoglobin polymerizes within red blood cells (RBCs), causing them to adopt an elongated “sickle” shape. Sickle RBCs damage vasculature, leading to severe symptoms, ultimately diminishing patient quality of life and reducing lifespan. Here, we use codelivery of a pre-formed Cas9 ribonucleoprotein complex (RNP) and a singlestranded DNA (ssDNA) oligonucleotide donor to drive sequence replacement at the SCD SNP in human CD34+ hematopoietic stem/progenitor cells (HSPCs). Corrected HSPCs from SCD patients produce less sickle hemoglobin protein and correspondingly increased wild-type hemoglobin when differentiated into erythroblasts. When injected into immunocompromised mice, treated HSPCs maintain editing long-term at therapeutically relevant levels. These results demonstrate that the Cas9 RNP/ssDNA donor approach can mediate efficient HSPC gene editing and could form the basis for treatment of SCD by autologous hematopoietic cell transplantation. ER -