Fig. 1.

Creating universal CAR T cells with genome editing. A Healthy donor T cells isolated from PB or UCB are genetically modified to express CAR. VL and VH chains are linked by a flexible peptide to form the scFv that recognizes tumor antigens. The hinge connects the scFv to the TM that anchors the receptor to the T cell’s membrane. TCR-derived CD3ζ and one or more co-stimulatory signaling domains activate CAR T cells. B To avoid alloreactivity, TCR-KO CAR T cells can be generated using genome editing techniques such as paired TALENs, composed of TALEs fused to FokI endonucleases for targeted DNA cleavage. C TCR KO can also be achieved using CRISPR-Cas9. Cas9 endonucleases and sgRNA form RNP complexes that cleave DNA at HNH and RuvC nuclease active sites. D DSBs from DNA cleavage are repaired via NHEJ or HDR mechanisms.