Genomic engineering of pluripotent cells

A method of generating genome-engineered hematopoietic lineage cells from human induced pluripotent stem cells (iPSCs), the method comprising:

• (i) initiating differentiation of human iPSCs toward hematopoietic lineage cells to obtain differentiating cells;
• and
• (ii) genetically engineering the differentiating cells by introducing into the differentiating cells:
• a targeted endonuclease, wherein (1) the targeted endonuclease targets a gene, (2) the gene encodes a protein of an MHC class I complex or a protein of an MHC class II complex, (3) the targeted endonuclease binds to a target sequence in the gene and introduces a double-strand break in the gene, thereby generating an in/del in the gene, and (4) the in/del disrupts the gene in the genome of the differentiating cells;
• (iii) differentiating the genetically engineered cells to obtain a population of genome-engineered hematopoietic lineage cells comprising in their genomes disruption of the gene;
• and
• (iv) sorting the population of genome-engineered hematopoietic lineage cells to obtain an population of genome-engineered hematopoietic lineage cells enriched for CD45+CD7+ cells:
• wherein the iPSCs are differentiated without formation of embryoid bodies;
• wherein the population of genome-engineered hematopoietic lineage cells enriched for CD45+CD7+ cells comprises T cells or T cell progenitors;
• and
• wherein the genome-engineered hematopoietic lineage cells enriched for CD45+CD7+ cells comprise (a) HLA-I deficiency, or (b) HLA-II deficiency, as compared to a corresponding iPSC-derived cell without the genetically engineering.