Apigenin Sensitizes Melanoma to Chemo/Radiotherapy and Potentiates Immune Killing by Reducing T Cell PD-1 Expression

Preclinical and clinical studies have suggested that combination therapies could potentially lead to further improved clinical outcomes of melanoma. This study aimed to explore the effects and mechanisms of apigenin combination with chemotherapy or radiotherapy in suppressing melanoma. Colony formation, immunofluorescence, flow cytometric analysis, and apoptosis assay were used to detect the effects of apigenin, DTIC, irradiation, or apigenin combined with DTIC or irradiation in melanoma cells. Western blot analysis and semi-quantitative PCR were used to measure the levels of PD-1 protein and PDCD1 mRNA. Cycloheximide chase assays, PS-341, and chloroquine treatment were used to explore the mechanisms of PD-1 downregulatory effects by apigenin. Killing assay was used to assess the killing effects of combination therapies against melanoma cells. In vivo experiments were generated to investigate the impact of combination therapy on tumor growth and immune infiltration. We found that combination treatment was more effective in inhibiting colony formation and Ki-67 expression in melanoma cells. Combination treatment caused stronger cell cycle arrest at the G2/M phases and enhanced cellular apoptosis than monotherapies. Further, we observed that apigenin promoted the degradation of PD-1 in T cells via the proteasomal pathway. Killing assays showed that the combination treatment demonstrated significantly enhanced T cell-mediated killing. In vivo experiments confirmed that combined treatment noticeably inhibited melanoma growth and enhanced T cell infiltration into melanoma tissues. Apigenin promoted the degradation of PD-1 in T cells via the proteasomal pathway, in addition to synergizing with chemo/radiotherapy to inhibit melanoma growth.