Targeted RNA Base Editing with Endogenous ADARs in T Cells

• Applicant: Professor Dr. Thorsten Stafforst
• Subject Area: Biochemistry ; Hematology, Oncology ; Immunology ; Cell Biology
• Term: since 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 428802479

RNA base editing has emerged as a powerful tool for programmable and reversible manipulation of genetic information. Building on our recent development of two novel strategies—CLUSTER and RESTORE—that recruit endogenous ADAR enzymes (double-stranded RNA-specific adenosine deaminase) for targeted A(denosine)-to-I(nosine) RNA base editing, this proposal aims to further advance the technology. Specifically, we seek to enhance the efficiency and precision of RNA base editing, establish its utility in T cells and CAR T cells (chimeric antigen receptor), and explore its potential to improve immune cell function through modulation of T cell signaling and metabolism. The project is divided into three work packages. In WP1, we aim to establish a library screening protocol to enhance the CLUSTER technology, e.g. to access CLUSTER guide RNA faster and that achieve higher editing efficiency. In WP2, we aim to establish targeted A-to-I RNA base editing in T and CAR T cells, including the delivery of RESTORE oligonucleotides by lipid-conjugation. In WP3, we aim to apply targeted RNA base editing to modulate CAR T cell function, e.g. to modulate T cell metabolism and to functionally characterize the effect of specific RNA base edits on clinically desirable CAR T cell features, like exhaustion. The overarching aim is to improve T cell fitness and counteract T cell exhaustion. The latter limits current T cell therapies and results in low T cell durability leading to relapse and/or a lack of efficiency in solid tumors. With this action, we will break new ground by running RNA base editing screens in T cells with the long-term vision to improve T cell therapies and other adaptable immunotherapies.

• DFG Programme: Research Grants