Effects of anti-PD-L1 monoclonal antibody and EGFR-TKI on the expression of PD-L1 and function of T lymphocytes in EGFR-mutated lung cancer cells

Abstract

Objective: To investigate the effects of anti-PD-L1 monoclonal antibody and EGFR-TKI on expression of soluble PD-L1 and function of T lymphocytes in EGFR-mutated lung cancer cells. Methods: Flow cytometry was used to analyze the expression of membrane PD-LI. ELISA was performed to detect the level of sPD-L1 in the supernatant of cultured EGFR-mutated and wild type lung cancer cells before and after erlotinib treatment.After treated with anti-PD-L1 monoclonal antibody alone and in combination with erlotinib, the proliferation of T lymphocytes in co-culture system was measured using Cell Counting Kit-8 (CCK-8) assay. The expression levels of PD-LI and IFN-γ in tumor cells and T lymphocytes treated with erlotinib in co-culture system were analyzed by flow cytometry and ELISA, respectively. Results: PD-L1 was highly expressed in EGFR-mutated lung cancer PC9 cells (78.7±3.1)% and HCC827 cells (82.7±2.6)%.After treated with erlotinib, the expression rates of membrane PD-L1 in PC9 and HCC827 cells were down-regulated (64.7%±3.1% and 73.0%±2.6%, respectively), significantly lower than that in the two cell lines without erlotinib treatment (P<0.05), and the expression levels of sPD-L1 in the supernatant of PC9 and HCC827 cells were also down-regulated (0.680±0.120)ng/ml and (0.903±0.047)ng/ml, respectively, significantly lower than that in the two cell lines without erlotinib treatment (P<0.01). However, no significant changes of membrane PD-L1 and sPD-L1 expression were found in EGFR wild type lung cancer cells (H1299 and A549) before and after erlotinib treatment. In the co-culture system composed of T cells and EGFR-mutated lung cancer cells, treatment with erlotinib alone promoted the proliferation of T lymphocytes (P<0.05), and combined treatment of anti-PD-L1 monoclonal antibody with erlotinib had a stronger effect (P<0.05). In the co-culture system composed of T cells and EGFR wild type cell lines, the proliferation of T cells was not changed after using erlotinib alone or combination of erlotinib and anti-PD-L1 monoclonal antibody (P>0.05). Before and after treatment with erlotinib, the secretion levels of IFN-γ were (856.0±70.3)pg/ml and (1 697.3±161.0)pg/ml, respectively, showing a significant difference (P<0.001). The expression rates of membrane PD-L1 were (76.2±0.5)% and (50.9±0.9)%, respectively, also showing a significant difference (P<0.001). However, no significant changes in the expression of IFN-γ and membrane PD-L1 were found in the co-culture system composed of T cells and A549 cells. Conclusions: Anti-PD-L1 monoclonal antibody combined with EGFR-TKI can effectively promote the proliferation and secretion function of T lymphocytes in the microenvironment of EGFR-mutated lung cancer cells.