{"id":"doi:10.1016/j.ymthe.2026.07.033","kind":"article","title":"A singular base-editing platform for polyfunctional multiplex engineering of immune cells","authors":["Skeate JG","Slipek NJ","Lahr WS","Roy S","Wick BJ","Kar B","Chang JW","Stelljes EM","Gilkey AK","Thenge PP","Diers MD","Krueger JB","Niemeyer EM","Lonetree CL","Kluesner MG","Bell JB","Clement K","Provenzano PP","Moriarity BS","Webber BR"],"author_count":20,"year":2026,"published":"2026-07-18","venue":"Molecular therapy : the journal of the American Society of Gene Therapy","cited_by":2,"doi":"10.1016/j.ymthe.2026.07.033","arxiv_id":null,"nct_id":null,"source_url":"https://doi.org/10.1016/j.ymthe.2026.07.033","archive_url":"https://space-kitty.com/papers/doi:10.1016/j.ymthe.2026.07.033","updated":null,"pmid":"42470099","pmcid":null,"openalex_id":null,"pdf_url":null,"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 knockin (KI), we expand the use of base editor nickase activity to stimulate homology-directed repair (HDR) and insert clinically relevant chimeric antigen receptors (CARs) into specific loci. Through a novel single-guide RNA design strategy and a DNA template delivered by a recombinant adeno-associated virus, 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/CD3E/PDCD1/CISH), we achieve single-step generation of highly functional off-the-shelf CAR T cell products with enhanced function. Importantly, we found no detectable translocations or significant off-target edits and demonstrated efficacy against multiple cancer lines and a suppressive 3D spheroid culture model. This efficient engineering process of \"Iterative Nicking for Synchronous Engineered Reprogramming of T cells\" (INSERT) establishes a safe, simplified platform for advanced therapeutic CAR T engineering.","abstract_license":"CC-BY (article license as recorded by Europe PMC; abstract reproduced unmodified apart from markup removal)","license":"CC-BY","is_open_access":false,"retracted":false,"peer_reviewed":null,"categories":[],"details":null,"provenance":"europepmc","attribution":"Abstract of \"A singular base-editing platform for polyfunctional multiplex engineering of immune cells\" by Skeate JG, Slipek NJ, Lahr WS et al., https://doi.org/10.1016/j.ymthe.2026.07.033, used under its Creative Commons Attribution license (CC-BY); markup removed, otherwise unmodified.","topics":[{"slug":"gene-editing-therapies","name":"CRISPR & Gene-Editing Therapies","domain_slug":"medicine"}],"cited_in_dossiers":[]}