CRISPR/Cas9-mediated PD-1 disruption enhances human mesothelin-targeted CAR T cell effector functions

Area of Science:

• Immunology
• Oncology
• Gene Editing

Background:

• Chimeric antigen receptor (CAR) T cells show promise for cancer therapy, but their efficacy against solid tumors is often limited by immune checkpoints like programmed cell death protein 1 (PD-1).
• Mesothelin is overexpressed in triple-negative breast cancer (TNBC), making it a potential target for CAR T cell therapy.

Purpose of the Study:

• To investigate the efficacy of genetically engineered CAR T cells with disrupted PD-1 for targeting mesothelin-expressing TNBC.
• To evaluate if CRISPR/Cas9-mediated PD-1 gene disruption enhances CAR T cell function and anti-tumor activity in vitro and in vivo.

Main Methods:

• Utilized CRISPR/Cas9 ribonucleoprotein-mediated gene editing to disrupt the PD-1 gene locus in human primary T cells.
• Generated mesothelin-targeted CAR T cells with PD-1 disruption.
• Assessed CAR T cell proliferation, cytokine production, and cytotoxicity against PD-L1-expressing cancer cells in vitro.
• Evaluated tumor control and relapse prevention in vivo using a TNBC mouse model.

Main Results:

• PD-1 gene disruption significantly reduced the PD-1-expressing T cell population without affecting CAR T cell proliferation.
• PD-1-disrupted CAR T cells exhibited augmented cytokine production and cytotoxicity against cancer cells in vitro.
• CAR T cells with PD-1 disruption demonstrated superior tumor control and relapse prevention in vivo compared to standard CAR T cells or those with antibody blockade.

Conclusions:

• CRISPR/Cas9-mediated PD-1 disruption is a viable strategy to enhance CAR T cell efficacy against solid tumors like TNBC.
• Integrated immune checkpoint blockade via gene editing offers a promising alternative CAR T cell therapy approach for improved adoptive transfer outcomes.