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Updated: Jun 4, 2025
Author Spotlight: Enhancing CAR-T Cell Function in Syngeneic Tumor Models
Published on: February 2, 2024
Leucine zipper-based immunomagnetic purification of CAR T cells displaying multiple receptors
Scott E James1,2,3,4,5,6, Sophia Chen7,8, Brandon D Ng9,7
1Adult Bone Marrow Transplantation Service, Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, NY, USA. scjames@coh.org.
Abstract:
Resistance to chimaeric antigen receptor (CAR) T cell therapy develops through multiple mechanisms, most notably antigen loss and tumour-induced immune suppression. It has been suggested that T cells expressing multiple CARs may overcome the resistance of tumours and that T cells expressing receptors that switch inhibitory immune-checkpoint signals into costimulatory signals may enhance the activity of the T cells in the tumour microenvironment. However, engineering multiple features into a single T cell product is difficult because of the transgene-packaging constraints of current gene-delivery vectors. Here we describe a cell-sorting method that leverages leucine zippers for the selective single-step immunomagnetic purification of cells co-transduced with two vectors. Such 'Zip sorting' facilitated the generation of T cells simultaneously expressing up to four CARs and coexpressing up to three 'switch' receptors. In syngeneic mouse models, T cells with multiple CARs and multiple switch receptors eliminated antigenically heterogeneous populations of leukaemia cells coexpressing multiple inhibitory ligands. By combining diverse therapeutic strategies, Zip-sorted multi-CAR multi-switch-receptor T cells can overcome multiple mechanisms of CAR T cell resistance.
Insights
This study introduces Zip sorting, a novel cell-sorting method enabling the creation of advanced T cells. These engineered T cells (chimaeric antigen receptor T cells) overcome resistance in cancer therapy.
Area of Science:
- Immunotherapy
- Cellular Engineering
- Cancer Research
Background:
- Chimaeric antigen receptor (CAR) T cell therapy faces resistance via antigen loss and immune suppression.
- Engineering T cells with multiple CARs or switch receptors is challenging due to vector limitations.
Purpose of the Study:
- To develop a method for generating T cells with multiple CARs and switch receptors.
- To overcome resistance mechanisms in CAR T cell therapy.
Main Methods:
- A novel cell-sorting method named 'Zip sorting' was developed using leucine zippers.
- This method allows for single-step immunomagnetic purification of co-transduced cells.
- Facilitated simultaneous expression of up to four CARs and three switch receptors.
Main Results:
- Zip sorting enabled the generation of T cells with multiple CARs and switch receptors.
- These engineered T cells effectively eliminated antigenically heterogeneous leukemia cells in mouse models.
- Demonstrated overcoming of multiple resistance mechanisms in CAR T cell therapy.
Conclusions:
- Zip-sorted multi-CAR multi-switch-receptor T cells represent a promising strategy to overcome CAR T cell resistance.
- This approach combines diverse therapeutic modalities to enhance anti-cancer activity.
- The Zip sorting method overcomes transgene-packaging constraints for complex T cell engineering.
