Using gene-disrupted allogeneic T cells as universal effector cells provides an alternative to and potentially improves current chimeric antigen receptor (CAR) T cell therapy against cancers and infectious diseases.
The CRISPR/Cas9 system has recently emerged as a simple and efficient way for multiplex genome engineering. By combining the lentiviral delivery of CAR and CRISPR RNA electroporation to co-introduce RNA encoding the Cas9 and gRNAs targeting endogenous TCR, beta-2 microglobulin (B2M) and PD1 simultaneously, to generate gene-disrupted allogeneic CAR T cells deficient of TCR, HLA class I molecule and PD1.
The CRISPR gene-edited CAR T cells showed potent anti-tumor activities, both in vitro and in animal models and were as potent as non-gene edited CAR T cells. In addition the TCR and HLA class I double deficient T cells had reduced alloreactivity and did not cause graft-versus-host disease. Finally, simultaneous triple genome editing by adding the disruption of PD1 led to enhanced in vivo anti-tumor activity of the gene-disrupted CART cells.
Gene-disrupted allogeneic CAR and TCR T cells could provide an alternative as a universal donor to autologous T cells, which carry difficulties and high production costs. Gene-disrupted CAR and TCR T cells with disabled checkpoint molecules may be potent effector cells against cancers and infectious diseases.