A Singular Base Editing Platform for Polyfunctional Multiplex Engineering of Immune Cells.

ABSTRACT: Current methods to engineer antigen-specific receptors rely on randomly integrating vectors or double-strand break induced targeted integration, both of which pose safety risks. To implement an all-in-one tool for multiplex knockout (KO) and knock in (KI), we expand the use of cytosine and adenine base editor (ABE) nickase activity to stimulate homology-directed repair (HDR) and insert clinically relevant chimeric antigen receptors (CARs) into specific loci. Through a novel sgRNA design strategy and a recombinant adeno-associated virus (rAAV) delivered DNA template, we enhanced the efficiency of ABE8e-stimulated HDR in human T cells. By combining KI of CD19-, CD33-, or mesothelin-targeting CARs with >95% quadplex gene KO (B2M/CD3ε/PDCD1/CISH), we achieve single-step generation of hi

SUBMITTER: Skeate JG 

PROVIDER: S-EPMC12338553 | biostudies-literature | 2025 Jul

REPOSITORIES: biostudies-literature