Long-term persistence and functionality of adoptively transferred antigen-specific T cells with genetically ablated PD-1 expression

Abstract

Engagement of the inhibitory T cell receptor programmed cell death protein 1 (PD-1) associates with dysfunctional states of pathogen- or tumor-specific T cells. Accordingly, systemic antibody-mediated blockade of PD-1 has become a central target for immunotherapies but is also associated with severe toxicities due to loss of peripheral tolerance. Therefore, selective ablation of PD-1 expression on adoptively transferred T cells through direct genetic knockout (KO) is currently being explored as an alternative therapeutic approach. However, since PD-1 might also be required for the regulation of physiological T cell function and differentiation, the suitability of PD-1 as an engineering target is controversial. In this study, we systematically investigated the maintenance of T cell functionality after CRISPR/Cas9-mediated PD-1 KO in vivo during and after acute and chronic antigen encounter. Under all tested conditions, PD-1 ablation preserved the persistence, differentiation, and memory formation of adoptively transferred receptor transgenic T cells. Functional PD-1 KO T cells expressing chimeric antigen receptors (CARs) targeting CD19 could be robustly detected for over 390 d in a syngeneic immunocompetent mouse model, in which constant antigen exposure was provided by continuous B cell renewal, representing the longest in vivo follow-up of CAR-T cells described to date. PD-1 KO CAR-T cells showed no evidence for malignant transformation during the entire observation period. Our data demonstrate that genetic ablation of PD-1 does not impair functionality and longevity of adoptively transferred T cells per se and therefore may be pursued more generally in engineered T cell-based immunotherapy to overcome a central immunosuppressive axis.

TIM-3, LAG-3, or 2B4 gene disruptions increase the anti-tumor response of engineered T cells

“…In a clinical trial with PD-1 disrupted cells edited for T cell specificity, the frequency of cells with edits in the PDCD1 locus decreased from 25% to 5% of the cells expressing the transgenic TCR at 4 months after infusion, consistently with mouse studies of chronic infection where PD-1 deficient T cells proved less able to establish memory ( 23 ). However, recent reports challenged this hypothesis and demonstrated in a murine model long-term persistence and function of murine adoptively transferred antigen-specific T cells with genetically ablated PD-1 expression ( 67 ).…”

Revolutionizing cancer treatment: enhancing CAR-T cell therapy with CRISPR/Cas9 gene editing technology

“…Given that blocking immune checkpoints has the potential to reactivate T cells, a key question of interest would be whether selective reduction or elimination of immune checkpoint genes on T cells effectively maintain or enhance CAR-T cell function? Research has proved that employing CRISPR/Cas9 to delete PD-1 resulted in enhanced long-term persistence and activity of CAR-T cells (preclinical study) ( 20 ), and similarly, the deletion of CTLA-4 using CRISPR/Cas9 improved the proliferation and activity of CAR-T cells (preclinical study) ( 21 ). Numerous investigations conducted thus far have demonstrated that the abrogation of immune checkpoint inhibition can effectively enhance and optimize the performance of CAR-T cells across various dimensions ( Table 2 ).…”

Gene editing technology to improve antitumor T-cell functions in adoptive immunotherapy

“…In addition to the epigenetic modification, antitumor functions of CAR-T cells can be potentiated through ablation of various genes. For example, PDCD1 [ 52 , 53 ], CTLA4 [ 54 ], TGFBRII [ 55 ], and A2AR knockout [ 56 ] can block the suppressive signals from the tumor microenvironment and enhance effector functions of CAR-T cells.…”