1. Generation of Allogeneic CAR T Cells through Specific Degradation of the T Cell Antigen Receptor by E3 Ubiquitin Ligase Fusion Proteins
- Author
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Michael J. Harris, Hao Chen, Tianyu Cai, Yuting Yi, Qiaowen Deng, Yi Yao, Tianle Lan, Yanfeng Guo, Xiufang Xu, Xian Wen, Joshua E. McGee, Daniella Tatang, James Brock, Feng Shi, and Li Zhou
- Subjects
Gene Editing ,T-Lymphocytes ,Ubiquitin-Protein Ligases ,Hematopoietic Stem Cell Transplantation ,Receptors, Antigen, T-Cell ,Biomedical Engineering ,General Medicine ,Biochemistry, Genetics and Molecular Biology (miscellaneous) - Abstract
Receptor downregulation is instrumental for many therapeutic interventions. Receptor knockout through gene-editing technologies is efficient but can introduce off-target mutations and chromothripsis. Regulation of gene expression at the protein level is a promising alternative. Here, we present results showing the targeted T cell antigen receptor (TCR) degradation using chimeric E3 fusion proteins that we call Receptor Targeting Chimeras (ReceptorTAC). We show that TCR degradation is dependent on enzymatically active, membrane-anchored E3 ligase variants. TCR specificity was achieved by direct fusion of an E3 domain to the CD3ζ transmembrane sequence. Jurkat and primary T cells stably expressing the ReceptorTAC constructs showed significantly reduced responses to TCR stimulation. We also used our ReceptorTAC technology to generate TCR-deficient, claudin18.2-specific CAR T cells, where the activity of the CAR was unaffected by the expression of the ReceptorTAC. These data indicate that our ReceptorTAC molecule can be used to generate allogeneic CAR T cells.
- Published
- 2022
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