Panoply™ Human HLA-G Knockdown Stable Cell Line

Immunotherapy for solid tumors has long been hampered by the development of an immunosuppressive tumor microenvironment and the lack of specific tumor-associated antigens that can be targeted across diverse solid tumor types. Human leukocyte antigen G (HLA-G), an immune checkpoint protein (ICP), is newly expressed on most tumor cells as a means of evading immune attack and has recently been demonstrated in vitro as a useful target for chimeric antigen receptor (CAR)-T therapy for leukemia. Here, researchers developed a novel CAR strategy using natural killer (NK) cells as effector cells, characterized by enhanced cytolysis through DAP12-based intracellular signaling. Results showed that HLA-G CAR-transduced NK cells exhibited potent cytolysis against breast, brain, pancreatic, and ovarian cancer cells in vitro and inhibited xenograft tumor growth and prolonged median survival in orthotopic mouse models. In tumor co-culture experiments, the anti-HLA-G scFv portion promoted Syk/Zap70 activation in NK cells, suggesting a potential reversal of HLA-G-mediated immunosuppression, thereby restoring the cytolytic function of natural NK cells. Low-dose chemotherapy further induced tumor HLA-G expression, and combined with anti-HLA-G CAR-NK therapy, it achieved extensive tumor ablation in vitro and in vivo. Tumor HLA-G upregulation involves inhibition of DNMT1 and demethylation of the transporter associated with antigen processing 1 promoter.

Here, researchers tested the antitumor activity of HLA-G CAR-NK cells using MDA-MB-231, DBTRG-05MG, AsPC-1, and SKOV3 cells as in vitro models. At the same E:T ratio, with increasing time and NK cell titers, anti-HLA-G CAR-NK cells exhibited enhanced cytotoxicity compared to parental control NK cells (Figure 1D). Using stably HLA-G knockdown MDA-MB-231 and AsPC-1 cells, researchers confirmed that the enhanced killing capacity of HLA-G CAR-NK cells was dependent on HLA-G expression, and these HLA-G knockdown cells were more sensitive to parental NK cell-mediated cytotoxicity (Figure 1E). Taken together, these results demonstrate that HLA-G CARs can enhance NK cell cytotoxicity against a variety of solid tumor cells and that this cytotoxicity is HLA-G dependent.

Figure 1. Cytotoxic killing of multiple solid tumor cell lines by anti-HLA-G CAR-NK cells. (Jan C I, et al., 2021)