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Updated: Aug 8, 2025
Manufacturing Chimeric Antigen Receptor CAR T Cells for Adoptive Immunotherapy
Published on: December 17, 2019
CD19 CAR antigen engagement mechanisms and affinity tuning
Changhao He1, Jorge Mansilla-Soto2,3, Nandish Khanra1
1Department of Physiology and Biophysics, Weill Cornell Medical College, New York, NY, USA.
Abstract:
Chimeric antigen receptor (CAR) T cell therapy relies on T cells that are guided by synthetic receptors to target and lyse cancer cells. CARs bind to cell surface antigens through an scFv (binder), the affinity of which is central to determining CAR T cell function and therapeutic success. CAR T cells targeting CD19 were the first to achieve marked clinical responses in patients with relapsed/refractory B cell malignancies and to be approved by the U.S. Food and Drug Administration (FDA). We report cryo-EM structures of CD19 antigen with the binder FMC63, which is used in four FDA-approved CAR T cell therapies (Kymriah, Yescarta, Tecartus, and Breyanzi), and the binder SJ25C1, which has also been used extensively in multiple clinical trials. We used these structures for molecular dynamics simulations, which guided creation of lower- or higher-affinity binders, and ultimately produced CAR T cells endowed with distinct tumor recognition sensitivities. The CAR T cells exhibited different antigen density requirements to trigger cytolysis and differed in their propensity to prompt trogocytosis upon contacting tumor cells. Our work shows how structural information can be applied to tune CAR T cell performance to specific target antigen densities.
Insights
Researchers optimized chimeric antigen receptor (CAR) T cell therapy by tuning binder affinity to CD19. This structural approach improved CAR T cell sensitivity and cancer cell targeting for better therapeutic outcomes.
Area of Science:
- Immunology
- Biochemistry
- Structural Biology
Background:
- Chimeric antigen receptor (CAR) T cell therapy is a promising cancer treatment that utilizes engineered T cells to target cancer cells.
- The efficacy of CAR T cell therapy is significantly influenced by the affinity of the single-chain variable fragment (scFv) binder, which recognizes specific cell surface antigens.
- CAR T cells targeting CD19 have shown remarkable success in treating B cell malignancies and are approved by the FDA.
Purpose of the Study:
- To elucidate the structural basis of CAR T cell function by determining the cryo-electron microscopy (cryo-EM) structures of CD19 in complex with key binders.
- To leverage structural insights and molecular dynamics simulations to engineer CAR T cell binders with modulated affinities.
- To evaluate how variations in binder affinity impact CAR T cell performance, including tumor recognition sensitivity, cytolysis induction, and trogocytosis.
Main Methods:
- Obtained cryo-EM structures of the CD19 antigen complexed with the FMC63 and SJ25C1 binders.
- Performed molecular dynamics simulations based on the obtained structures.
- Engineered novel binders with altered affinities (lower and higher) based on simulation data.
- Assessed the functional performance of CAR T cells engineered with these modified binders in vitro.
Main Results:
- Determined high-resolution cryo-EM structures of CD19 bound to FMC63 and SJ25C1.
- Successfully designed and created CAR T cell binders with precisely tuned affinities.
- Demonstrated that CAR T cells with altered binder affinities exhibit distinct sensitivities to target antigen density.
- Observed differential induction of cytolysis and trogocytosis based on CAR T cell binder affinity and target cell antigen expression.
Conclusions:
- Structural information of CAR binders and their target antigens is crucial for understanding and optimizing CAR T cell function.
- Binder affinity can be rationally modulated using structural and computational approaches to fine-tune CAR T cell therapeutic potential.
- This study provides a framework for engineering CAR T cells with tailored specificities for diverse cancer antigen expression levels.
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