EF1α, rather than CMV promoter, is suitable for luciferase tag expression in target cells for <i>in vitro</i> cytotoxicity assays of CAR-T cells.

ABSTRACT: Precise assessment of the cytotoxic activity of engineered immune cell therapeutics, such as chimeric antigen receptor-engineered T (CAR-T) cells, is essential for their development and quality control. However, luciferase (Luc)-based viability assays, which evaluate target cell viability by overexpressing Luc tags and measuring chemiluminescent signals, may yield biased results depending on the promoter driving Luc expression. This study demonstrates that CAR-T cells can enhance cytomegalovirus (CMV) promoter-driven transcription in target cells via the interferon-gamma (IFN-γ)/nuclear factor κB (NF-κB) signaling pathway, leading to elevated Luc expression and a discrepancy between chemiluminescent signals and actual target cell death. These findings underscore the limitations of CMV prom

SUBMITTER: Hou R 

PROVIDER: S-EPMC12329255 | biostudies-literature | 2025 Sep

REPOSITORIES: biostudies-literature