Establishment of the Method for Screening Small Molecule Inhibitors Blocking the Interaction Between PD-1 and Its Ligand PD-L1

To screen the small molecule inhibitors blocking the interaction of PD-1 and PD-L1, the plasmids encoding GST-PD-L1 (AA26-236) and His-Trx-PD-1 (AA21-163) were constructed, and the fusion proteins were expressed in E. coli BL21-CodonPlus (DE3)-RIL cells and purified. GST pull-down experiment indicated that PD-1 directly interacts with PD-L1. The screening model of small molecule inhibitors blocking PD-1/PD-L1 interaction using competitive ELISA assay was established, and two small molecule chemicals named S2468 (Fenbendazole) and S2084 (Duloxetine HCl) were screened with inhibition rate 0.49 and 0.52. While S2485 (Mitoxantrone Hydrochloride) and S1957 (Sulfamethizole) were also screened with inhibition rate 1.8 and 1.6, which could increase the binding between PD-1 and PD-L1.