CRISPR/Cas9-Mediated Knockout of DGK Improves Antitumor Activities of Human T Cells

Area of Science:

• Immunology
• Cancer Biology
• Gene Editing

Background:

• T-cell therapy efficacy is limited by tumor-induced immunosuppression.
• Current strategies like PD-1/CTLA-4 blockade show limited clinical success.
• Targeting T-cell signaling pathways is crucial for enhancing anticancer immunity.

Purpose of the Study:

• To investigate the potential of diacylglycerol kinase (DGK) knockout to enhance T-cell function.
• To evaluate the impact of DGK ablation on CAR-T cell effector functions and resistance to immunosuppression.
• To assess the therapeutic efficacy of DGK-knockout CAR-T cells in a glioblastoma xenograft model.

Main Methods:

• Utilized CRISPR/Cas9 gene editing to knock out diacylglycerol kinase (DGK) in CAR-T cells.
• Assessed augmented T-cell receptor (TCR) signaling and effector functions in vitro.
• Evaluated CAR-T cell resistance to immunosuppressive factors (TGFβ, PGE2) and repeated tumor stimulation.
• Tested efficacy in a U87MGvIII glioblastoma xenograft mouse model.

Main Results:

• DGK knockout significantly enhanced CAR-T cell effector functions via increased TCR signaling.
• DGK-ablated CAR-T cells demonstrated resistance to immunosuppressive factors and sustained function under repeated stimulation.
• Significant regression of glioblastoma tumors was observed in the xenograft mouse model.
• CRISPR/Cas9-mediated DGK knockout improved antitumor immunity in vivo.

Conclusions:

• CRISPR/Cas9-mediated knockout of DGK effectively enhances CAR-T cell effector functions.
• DGK ablation offers a promising strategy to overcome tumor immunosuppression in T-cell therapy.
• This approach holds potential for treating solid cancers, including glioblastoma.