Turning "trashed" genomic loci into treasurable sites for integrating chimeric antigen receptors in T and NK cells.

Abstract

Chimeric antigen receptor (CAR)-based immune cell therapy involves genetically engineering immune cells, such as T cells and natural killer (NK) cells, to express CARs that can specifically recognize target antigens. This modification enables T/NK cells to selectively eliminate tumor cells following adoptive transfer. One common approach to stably integrate CARs into the genome of T/NK cells is through retroviral or lentiviral vectors. However, these vectors mediate semi-random gene integration, posing risks such as oncogenic mutations, gene silencing, and variable CAR expression levels. Targeted integration of CAR genes into the specific genomic locus could overcome these limitations, but identifying the optimal integration sites to maximize the safety and efficacy of CAR-T/NK cell products remains a critical question. Improper integration sites may disturb the endogenous genes surrounding the integration sites, raising safety concerns. Additionally, regulatory elements at the integration sites, such as promoters, can influence the expression level of CAR genes, thus affecting the efficacy of CAR-T/NK cells. In this review, we summarized current strategies for selecting integration sites and promoters in the engineering of CAR-T/NK cells to achieve potent anti-tumor efficacy in preclinical studies.