Affinity-tuned mesothelin CAR T cells demonstrate enhanced targeting specificity and reduced off-tumor toxicity

Area of Science:

• Immunology
• Oncology
• Biotechnology

Background:

• Chimeric antigen receptor (CAR) T cell therapy shows promise for solid tumors but is limited by severe toxicities.
• On-target, off-tumor toxicity occurs when CAR T cells attack healthy tissues expressing low levels of the target antigen.
• Mesothelin (MSLN) is a promising target for cancers like mesothelioma and ovarian cancer, but its expression on healthy mesothelial cells poses a toxicity risk.

Purpose of the Study:

• To investigate the impact of CAR T cell affinity for MSLN on tumor response and toxicity.
• To develop affinity-tuned CAR T cells for improved safety and efficacy in MSLN-expressing solid tumors.
• To evaluate the preclinical efficacy and safety of MSLN-targeting CAR T cells with varying affinities in mouse models.

Main Methods:

• Construction of CARs utilizing mutant single-domain nanobodies targeting MSLN with a broad range of affinities (nanomolar to micromolar KD).
• Evaluation of CAR T cell expansion, tumor infiltration, and systemic toxicity in mouse models with MSLN-expressing tumors.
• Comparative analysis of high-affinity versus affinity-tuned CAR T cells in preclinical settings.

Main Results:

• High-affinity MSLN CAR T cells (low nanomolar KD) showed extensive systemic expansion but minimal tumor infiltration and were associated with fatal off-tumor toxicity.
• Reducing CAR T cell affinity towards micromolar KD resulted in restricted tumor infiltration and improved selectivity for MSLN-high tumors.
• Affinity tuning of MSLN CAR T cells demonstrated a correlation between decreased affinity and enhanced tumor-specific targeting.

Conclusions:

• High-affinity MSLN-targeting CAR T cells can cause life-threatening on-target, off-tumor toxicity.
• Affinity modulation of CAR T cells is a viable strategy to enhance selectivity for MSLN-high tumors and mitigate toxicity.
• These findings support the development of affinity-tuned CAR T cell therapies for safer and more effective treatment of MSLN-expressing solid tumors.