Abstract: Objective The objective of this study was to investigate the expression of interleukin-38(IL-38)in patients with breast cancer and its regulatory function to CD8⁺T cell activity. Methods 44 patients with breast cancer,25 patients with benign breast tumor,and 20 controls,who were treated in the First Affiliated Hospital of Xinxiang Medical University from July 2020 and September 2022.Mononuclear cells from plasma and peripheral blood of all subjects were isolated,tumor-infiltrating lymphocytes from tumor tissues of breast cancer patients were isolated,and CD8⁺T cells were purified.IL-38 protein level in the plasma was measured by enzyme-linked immunosorbent assay(ELISA).The relative level of IL-38 mRNA in the tissue was semi-quantified by real-time quantitative PCR.Recombinant human IL-38 was used to stimulate CD8⁺T cells from peripheral blood and tumor tissue from patients with breast cancer.A co-culture system was established between CD8⁺T cells and breast cancer MCF-7 cell line.The percentage of target cell death was calculated by measuring lactate dehydrogenase level in the supernatants.The levels of perforin,granzyme B,interferon-γ and tumor necrosis factor-α(TNF-α)in the supernatants were measured by ELISA.The immune checkpoint molecules expression in CD8⁺T cells were detected by flow cytometry. Results The levels of plasma IL-38 were significantly higher in patients with breast cancer(74.23±19.88 pg/mL)compared with in patients with benign breast tumor(62.87±16.27 pg/mL,P=0.018)and controls(61.77±12.75 pg/mL,P=0.013).The relative expression of IL-38 mRNA in tumor tissues was significantly higher than in para-tumor tissues(1.57±0.22 vs. 1.00±0.18,P＜0.001).The proportion of target cell death induced by peripheral and tumor-infiltrating CD8⁺T cells,and the levels of perforin and granzyme B secretion in direct contact co-culture group were higher than those in indirect contact co-culture group(P＜0.05).There were no significant differences of either interferon-γ or TNF-α levels between direct contact and indirect contact co-culture group(P＞0.05).In the direct contact co-culture group,the levels of target cell death proportion,perforin,granzyme B,interferon-γ and TNF-α l in the IL-38 stimulation group were lower than those in the non-stimulation group(P＜0.05).In the indirect contact co-culture group,the target of cell death proportion,interferon-γ and TNF-α the IL-38 stimulation group were also lower than those in the non-stimulation group(P＜0.05).However,there were no statistical differences of either perforin or granzyme B levels between the IL-38 stimulation group and non-stimulation group within the indirect contact co-culture group(P＞0.05).There were also no differences in the levels of immune checkpoint molecules in CD8⁺T cells between the non-stimulation group and the IL-38 stimulation group(P＞0.05). Conclusion Highly expressed IL-38 in patients with breast cancer may be involved in inducing CD8⁺T cell functional failure.

Key words: Breast cancer, Interleukin-38, CD8⁺T-lymphocytes, Anti-tumor