T Cell Activators Exhibit Distinct Downstream Effects on Chimeric Antigen Receptor T Cell Phenotype and Function

Area of Science:

• Immunology
• Biotechnology
• Cell Therapy

Background:

• T cell activation is crucial for chimeric antigen receptor (CAR) T cell manufacturing.
• Activator reagents trigger T cell receptor (TCR) and provide costimulation, influencing CAR T cell culture attributes.

Purpose of the Study:

• To explore various T cell activation reagents and their effects on CAR T cell attributes.
• To examine how different delivery mechanisms of anti-CD3/anti-CD28 impact CAR T cell activation, expansion, gene expression, and function.

Main Methods:

• Four distinct activators were tested: Dynabeads, TransAct, Cloudz, and Microbubbles, all using anti-CD3 and anti-CD28.
• Cells were transduced with a lentiviral vector encoding a bivalent CD19/CD22 CAR.
• Cell counts and flow cytometry monitored cultures; functional assays assessed target cell killing and cytokine secretion.

Main Results:

• Cloudz activator skewed the CD4/CD8 ratio towards CD8 T cells.
• Dynabeads stimulation yielded the highest naive T cell subset, while Cloudz resulted in the most effector/effector memory cells.
• Cloudz-stimulated cells exhibited the highest target cell killing and proinflammatory cytokine secretion.

Conclusions:

• The method of presenting stimulatory antibodies to T cells influences CAR T cell activation and function.
• Different T cell activation reagents lead to distinct CAR T cell product attributes.
• These findings are critical for optimizing CAR T cell manufacturing for clinical applications.