Panoply™ Human CD274 Over-expressing Stable Cell Line

Programmed death ligand-1 (PD-L1 or CD274) is an important immune checkpoint molecule in the tumor microenvironment and an important target for cancer immunotherapy. It is well known that its expression in cancer cells can inhibit T cell-mediated antitumor responses, and this mechanism of action has become a target for cancer immunotherapy. Here, the researchers explored the effects of PD-L1 intervention on the cell biological functions of human esophageal cancer cell line Eca-109 cells, especially epithelial-mesenchymal transition (EMT). Clinical and pathological data showed that PD-L1 expression was higher in EMT-positive subgroup tumor samples than in EMT-negative subgroups. By knocking out or overexpressing wild-type and cytoplasmic domain truncation mutants to regulate PD-L1 expression in Eca-109 cells, the researchers confirmed that PD-L1 expression significantly promoted cell viability, migration, and EMT phenotypes. Furthermore, these studies also demonstrated that PD-1 fusion protein-mediated stimulation of PD-L1 and its cytoplasmic domain plays a key role in promoting the EMT phenotype of Eca-109 cells, suggesting that the PD-1 receptor often affects the regulation of PD-L1-mediated responses in esophageal cancer cells by triggering reverse signaling.

Here, the researchers tested whether the binding of PD-1 receptor to PD-L1 would affect PD-L1-mediated EMT promotion in ECA-109 cells. PD-L1 overexpressing ECA-109 cells were stimulated with different concentrations of PD-1 fusion protein for 24 hours. As shown in Figure 1A and B, the expression of E-cadherin in cells treated with 1 and 5 µg/ml PD-1 fusion protein was significantly reduced compared with the untreated group (blank group) or the low concentration group. However, the expression levels of N-cadherin and Zeb1 proteins were significantly increased under stimulation with PD-1 at both concentrations (1 and 5 µg/ml). Notably, the expression of vimentin was significantly increased by the three different concentrations of PD-1 fusion protein. In addition, changes in the mRNA expression levels of E-cadherin, N-cadherin, Zeb-1 and Vimentin genes in PD-L1 over-expressing Eca-109 cells treated with different concentrations of PD-1 fusion protein for 24 hrs were analyzed by RT-PCR. As shown in Figure 1C, the mRNA expression pattern was consistent with the expected changes observed in Figures 1A and B.

Figure 1. PD-1 fusion protein treatment promotes EMT phenotype in PD-L1 over-expressing Eca-109 cells. (Chen L, et al., 2017)