The influence of aranoza drug formulations and «empty» liposomes on the expression of PD-L1 и PD-L2 in human melanoma cell lines

Abstract

Introduction. In recent years, significant progress has been achieved in immunotherapy of tumors, however, little is known about the crosstalk between the tumor and the immune system. It seems reasonable to rise the question of patients treatment with PD-1 and its ligands blockers after chemotherapy. Objective. To investigate the expression of cell surface molecules PD-L1 and PD-L2 both at the mRNA and protein levels in human melanoma cell lines after exposure to «Aranoza, lyophilisate for preparation of solution for injections» (aranoza-lio), liposomal formulation of aranoza and «empty» liposomes without aranoza. Materials and methods. In this study we used 11 melanoma cell lines, 5 of which carried the BRAF mutation. In quantitative polymerase chain reaction in real time we investigated the level of PD-L1 and PD-L2 gene expression. Using flow cytometry we evaluated the expression of cell surface antigens PD-L1 and PD-L2. Results. PD-L1 and PD-L2 mRNA are expressed at a lower level in cells with BRAF mutations, differences were significant only for PD-L2 (p = 0.1373 andp = 0.0207respectively). A high basal level of protein PD-L1 and PD-L2 expression was observed in wild type BRAF melanoma cells. After exposure to liposomal aranoza the level of PD-L1 and PDL-2 mRNA expression was significantly reduced (p = 0.0004 and p = 0.0442 respectively) in compare to control non-treated cells. It is of interest, aranoza-lio and «empty» liposomes increased the expression of PD-L1 (p <0.0001 and p = 0.0005 respectively) and PDL2 (p = 0.0005 and p = 0.0025). Of note, we observed an opposite effect in some melanoma cell lines. Moreover, changes in the expression of proteins PD-L1 and PD-L2 did not correlate with the mRNA level. The expression of PD-L1 protein after incubation with the liposomal aranoza increased dramatically compared to non-treated cells (p = 0.0269). Aranoza-lio and «empty» liposomes decreased the expression of PD-L1 protein (p = 0.0663 and p = 0.7213 respectively). The protein level of PD-L2 after exposure to liposomal aranoza did not change significantly (p = 0.1141), however, aranoza-lio and «empty» liposomes reduced the expression of PD-L2 (p = 0.0021 andp = 0.008). Conclusion. These findings are consistent with those of other researchers and confirm that the level of PD-L1 and PD-L2 mRNA and protein are modulated after treatment. Also, various drug formulations of Aranoza, as well as the «empty» liposomes, have different effects on the expression of PD-L1 and PD-L2 mRNA and proteins.