Massively parallel base editing to map variant effects in human hematopoiesis

Systematic evaluation of the impact of genetic variants is a critical need in the study and treatment of human physiology and disease. While specific mutations can be introduced by genome engineering, we still lack scalable approaches that are applicable to the important setting of primary cells, such as blood and immune cells. Here, we describe the development of massively parallel base-editing screens in human hematopoietic stem and progenitor cells. Such approaches enable functional screens for variant effects across any hematopoietic differentiation state. Moreover, they allow rich phenotyping through single-cell RNA sequencing readouts and careful characterization of editing outcomes through pooled single-cell genotyping. We apply the approach to several important problems, including designing improved leukemia immunotherapies, identifying non-coding variants modulating the expression of fetal hemoglobin, defining mechanisms regulating hematopoietic differentiation, and probing the pathogenicity of uncharacterized disease-associated variants. This approach will advance variant-to-function mapping in human hematopoiesis to identify the causes of diverse diseases.