Homo sapiens Targeted loci cultured

B cells differentiate into long-lived plasma cells that provide humoral immunity by secreting large quantities of antibodies. The ability to engineer primary human B cells to secrete a de novo protein may allow the creation of novel plasma cell therapies for protein deficiency diseases and other clinical applications. To achieve this goal, we developed methods for efficient genome editing of primary B cells isolated from peripheral blood, followed by ex vivo differentiation into plasma cells. By delivering CRISPR/Cas9 ribonucleoprotein (RNP) complexes under conditions of rapid B cell expansion, we achieved site-specific gene disruption at multiple loci in primary human B cells (with editing rates up to 94%). We first used this method to modulate plasma cell differentiation by disrupting key developmental regulatory genes. Next, we co-delivered RNPs with either single-stranded DNA oligonucleotide and achieved targeted sequence integration at high efficiency (up to 40%) via homology-directed repair. Our results highlight the utility of genome editing in studying human B cell biology and demonstrate a novel strategy for modifying human plasma cells to secrete therapeutic proteins.