Language
English
Publication Date
6-20-2024
Journal
Blood
DOI
10.1182/blood.2023020973
PMID
38493479
PMCID
PMC11196866
PubMedCentral® Posted Date
3-19-2024
PubMedCentral® Full Text Version
Post-print
Abstract
Chimeric antigen receptor (CAR)-redirected immune cells hold significant therapeutic potential for oncology, autoimmune diseases, transplant medicine, and infections. All approved CAR-T therapies rely on personalized manufacturing using undirected viral gene transfer, which results in nonphysiological regulation of CAR-signaling and limits their accessibility due to logistical challenges, high costs and biosafety requirements. Random gene transfer modalities pose a risk of malignant transformation by insertional mutagenesis. Here, we propose a novel approach utilizing CRISPR-Cas gene editing to redirect T cells and natural killer (NK) cells with CARs. By transferring shorter, truncated CAR-transgenes lacking a main activation domain into the human CD3ζ (CD247) gene, functional CAR fusion-genes are generated that exploit the endogenous CD3ζ gene as the CAR's activation domain. Repurposing this T/NK-cell lineage gene facilitated physiological regulation of CAR expression and redirection of various immune cell types, including conventional T cells, TCRγ/δ T cells, regulatory T cells, and NK cells. In T cells, CD3ζ in-frame fusion eliminated TCR surface expression, reducing the risk of graft-versus-host disease in allogeneic off-the-shelf settings. CD3ζ-CD19-CAR-T cells exhibited comparable leukemia control to TCRα chain constant (TRAC)-replaced and lentivirus-transduced CAR-T cells in vivo. Tuning of CD3ζ-CAR-expression levels significantly improved the in vivo efficacy. Notably, CD3ζ gene editing enabled redirection of NK cells without impairing their canonical functions. Thus, CD3ζ gene editing is a promising platform for the development of allogeneic off-the-shelf cell therapies using redirected killer lymphocytes.
Keywords
Killer Cells, Natural, Humans, CD3 Complex, Receptors, Chimeric Antigen, Animals, Mice, T-Lymphocytes, Cytotoxicity, Immunologic, Immunotherapy, Adoptive, Gene Editing, CRISPR-Cas Systems, Mice, Inbred NOD
Published Open-Access
yes
Recommended Citation
Kath, Jonas; Franke, Clemens; Drosdek, Vanessa; et al., "Integration of ζ-Deficient CARs Into the CD3ζ Gene Conveys Potent Cytotoxicity in T and NK Cells" (2024). Faculty, Staff and Students Publications. 6383.
https://digitalcommons.library.tmc.edu/baylor_docs/6383
Graphical Abstract