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Structure-function relationships of chimeric antigen receptors in acute T cell responses to antigen
Han Xu1, Agnes E Hamburger1, Jee-Young Mock1
1A2 Biotherapeutics, 30301 Agoura Rd., Agoura Hills, CA, 91301, United States.
Abstract:
Chimeric antigen receptors (CARs) and their parent signaling molecule, the T cell receptor (TCR), are fascinating proteins of increasing relevance to disease therapy. Here we use a collection of 1221 pMHC-directed CAR constructs representing 10 pMHC targets to study aspects of CAR structure-activity relationships (SAR), with particular focus on the extracellular and transmembrane structural components. These experiments that involve pMHC targets whose number/cell can be manipulated by peptide dosing in vitro enable systematic analysis of the SAR of CARs in carefully controlled experimental situations (Harris and Kranz, 2016). We find that CARs tolerate a wide range of structural variation, with the ligand-binding domains (LBDs) dominating the SAR of CAR antigen sensitivity. Notwithstanding the critical role of the LBD, CAR antigen-binding on the cell surface, measured by pMHC tetramer staining, is not an effective predictor of functional sensitivity. These results have important implications for the design and testing of CARs aimed toward the clinic.
Insights
Chimeric antigen receptors (CARs) tolerate structural changes, with ligand-binding domains (LBDs) key to antigen sensitivity. However, cell surface binding doesn't predict functional sensitivity, impacting CAR therapy design.
Area of Science:
- Immunology
- Molecular Biology
- Biotechnology
Background:
- Chimeric antigen receptors (CARs) and T cell receptors (TCRs) are crucial in disease therapy.
- Understanding CAR structure-activity relationships (SAR) is vital for optimizing therapeutic efficacy.
Purpose of the Study:
- To investigate CAR SAR, focusing on extracellular and transmembrane structural components.
- To analyze how structural variations affect CAR antigen sensitivity and function.
- To determine predictors of CAR functional sensitivity for clinical applications.
Main Methods:
- Systematic analysis of 1221 pMHC-directed CAR constructs targeting 10 pMHCs.
- In vitro experiments manipulating pMHC target number via peptide dosing.
- Measurement of CAR antigen-binding using pMHC tetramer staining.
Main Results:
- CARs exhibit tolerance to substantial structural variations.
- Ligand-binding domains (LBDs) are the primary determinants of CAR antigen sensitivity.
- Cell surface antigen-binding does not reliably predict functional CAR sensitivity.
Conclusions:
- CAR structural diversity is permissible, with LBDs playing a dominant role in antigen recognition.
- pMHC tetramer staining is an insufficient predictor of CAR functional sensitivity.
- Findings necessitate revised strategies for CAR design and preclinical testing in clinical settings.
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