Investigate the killing effect of NK cells treated by CD137 against breast cancer cells mediated by antibody dependent cell cytotoxicity (ADCC) .

Groups in phenotype and cytokine detection experiments: negative control group (NK cells not treated with human CD137 antibody) , CD137 antibody treatment group (NK cells treated with 10 μg/mL human CD137 antibody for 4 h) . Groups in cytotoxicity experiment: there are 8 groups according to the addition of NK cells, human CD137 antibody and cetuximab in the system. The ratio of CD16⁺ NK cell was detected by flow cytometry, the concentration of IFN-γ and TNF-α in the supernatant of NK cell culture media was detected by enzyme linked immunosorbent assay (ELISA) , and the cytotoxicity of NK cells against MDA-MB-231 and MDA-MB-453 cell lines mediated by cetuximab was measured by lactate dehydrogenase (LDH) . t test or factorial analysis was applied for statistical analysis.

Compared with NC group, the ratio of CD16⁺ NK cells and the concentration of IFN-γ and TNF-α in the CD137 antibody treatment group was significantly higher than that of the control group (79.57﹪± 0.92﹪vs 90.43﹪± 0.67﹪, 388.90 pg/mL ± 7.02 pg/mL vs 523.90 pg/mL±1.90 pg/ mL, 20.59 pg/mL±4.09 pg/mL vs 47.22 pg/mL±2.14 pg/mL, P < 0.05) . The concentration of IFN-γ and TNF-α secreted from CD137 antibody treated NK cells was (523.90±1.90) pg/mL and (47.22±2.14) pg/mL, higher than that of the control group (388.90±7.02) pg/mL (P < 0.01) and (20.59±4.09) pg/mL (P < 0.05) respectively. The results of three factor factorial analysis showed that NK cells, human CD137 antibody and cetuximab could kill MDA-MB-231 cells (F = 5227.276, 201.473, 1792.242, P < 0.001) , and the interaction of the three factors also had effect on the killing of MDA-MB-231 cells (F = 183.903, 1517.187, 33.483, P < 0.001) . The secondary interaction of the three factors was also statistically significant (F = 41.505, P < 0.001) .

All above, we have presented that CD137 antibody can enhance the secretion ability of NK cells to secrete IFN-γ and TNF-α. Meanwhile, it could upregulate the expression of CD16 molecules on the surface of NK cells, so that NK cells may enhance the cytotoxicity against epidermal growth factor receptor (EGFR) positive breast cancer cells through ADCC mediated by cetuximab.