Hyperoside exerts its anti-tumor activity by reducing the PD-L1 level in non-small cell lung cancer

Blocking the binding of programmed death 1 (PD-1) on the T cells and programmed death ligand 1 (PD-L1) on the tumor cells has become a hotspot in the field of tumor immunotherapy. Small-molecule checkpoint inhibitor targeting PD-1/PD-L1 axis is the new direction of tumor immunotherapy. In the present study, we investigated the anti-tumor role of hyperoside by regulating the PD-L1 level in non-small cell lung cancer (NSCLC). Changes of total PD-L1 and membrane PD-L1 levels were determined by Western blot, flow cytometry, and PD-1/PD-L1 interaction assays. The expression of mRNA level of PD-L1 was detected by real-time PCR. The cytotoxicity of activated human T cells toward co-cultured tumor cells was measured by cell impedance assay and crystal violet experiment. The antitumor effect of hyperoside in vivo was examined by C57BL/6 mice bearing Lewis xenograft tumor. Western blot and flow cytometry assay showed that hyperoside significantly downregulated the abundance of PD-L1 in H1975 and HCC827 cells in dose- and time-dependent manner. PD-1/PD-L1 binding assay revealed that hyperoside reduced the binding of tumor cells to recombinant PD-1 protein. In addition, hyperoside decreased the abundance of c-Myc, a key transcriptional regulator of PD-L1, in H1975 and HCC827 cells. Cell impedance and crystal violet staining indicated that hyperoside enhanced the killing activity of co-cultured T cells toward tumor cells. Animal experiments (all animal experiments were conducted in accordance with the Animal Ethics Committee of the Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences) revealed that hyperoside treatment displayed significant suppression in the growth of Lewis tumor xenografts in C57BL/6 mice with an inhibition rate of 48.3% at 25 mg·kg^-1. Our results demonstrate that hyperoside exerts its anti-NSCLC activity by reducing the PD-L1 level. Our study provides an important material basis and scientific basis for developing hyperoside into a new small molecule drug for tumor immunotherapy.