CRISPR-Cas9 disruption of PD-1 enhances activity of universal EGFRvIII CAR T cells in a preclinical model of human glioblastoma

摘要

Multiplexed CRISPR-Cas9 gene-editing is efficient in primary human T cells. Schematic representation of the EGFRvIII targeted CAR construct. Primary human T cells were stimulated, RNP electroporated and transduced to produce CAR T cells. Following expansion, cells were subjected to flow cytometry for TCR and B2M expression. Bivariate plot displays frequency of cells with both TCR and B2M deletion. EGFRvIII CAR T cells that have been gene-edited for PD-1 (CART-EGFRvIIIΔPD-1) do not have the ability to interact with PD-L1 expressed on target cells. Effector cells were incubated with irradiated U87vIII for 1 week and subjected to flow cytometric analysis for surface PD-1 expression. The control group contains cells gene-edited for both TCR and B2M, and mock transduced with AAV