ArticleJournal of translational medicine2026
Engineered PD1-NKG2D Dual-CAR NK92 cells broaden antitumor target recognition in preclinical tumor models.
Article in Journal of translational medicine, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
backgroundTo address challenges such as the complex manufacturing of CAR-T and the immunosuppressive tumor microenvironment (TME), CAR-NK cells offer greater potential as an "off-the-shelf" therapy. To broaden tumor recognition and reduce the risk of immune escape associated with single-target approaches, we developed a single-promoter-driven multicistronic CAR-NK92 system that employs NKG2D for broad recognition of stress ligands and combines a PD1-CAR to reverse PD-L1 inhibitory signaling, thereby significantly enhancing antitumor efficacy.
methodsA multicistronic construct co-expressing PD1-CAR and NKG2D-CAR was generated using a P2A peptide under the control of a single CMV promoter and introduced into NK92 cells. The expression of PD-L1 and MICA/B was screened across multiple tumor cell lines, and the functional robustness of PN-CAR-NK92 cells was evaluated in all models through in vitro cytotoxicity and cytokine secretion assays. The in vivo translational efficacy was further validated using an H1299 xenograft model, with a direct comparison between PN-CAR-NK92 cells and NK92 cells.
resultsThe multicistronic design enabled stable surface co-expression of both receptor modules, providing a structural basis for dual-target functionality. In vitro cytotoxicity assays demonstrated that PN-CAR-NK92 cells maintained robust antitumor activity across tumor cell lines with distinct PD-L1 and MICA/B expression profiles, whereas single-target CAR-NK92 cells displayed more restricted target specificity. These findings suggest that dual-target CAR engineering broadens antigen recognition coverage and may help reduce the limitations associated with single-target antigen dependence. Furthermore, PN-CAR-NK92 cells demonstrated significantly enhanced tumor suppression in the H1299 xenograft model compared with control groups.
conclusionsDual-target PD1/NKG2D CAR-NK92 cells exhibit broadened antitumor activity across tumor cells with distinct ligand-expression profiles and may represent a promising strategy to reduce the limitations associated with single-target CAR therapies.
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