The Research Of CD47’s Expression And Targeted Therapy On Ovarian Cancer In Vitro

Objective (1) To evaluate the expression of CD47in ovarian cancer, and analyze its relationship with clinicopathological parameters, and provide a theoretical basis for CD47monoclonal antibody targeted therapy in ovarian cancer.(2) To explore the treatment of using anti-CD47to promote the phagocytosis in vitro in ovarian cancer.(3) Preliminary explore the expression of CD47in ALDH+/ALDH-ovarian cancer cells, and provide an experimental basis for the research of ovarian cancer stem cells.Methods①Detected the expression of CD47in human ovarian cancer cell line SKOV-3, normal ovarian tissue cells, and ovarian cancer cells by flow cytometry, clarify its relationship with clinicopathological parameters in patients with ovarian cancer.②Analyze of mouse macrophages, human peripheral blood macrophages and human cord blood macrophages activity by cell morphology, CD14expression, and chicken erythrocytes engulfed experiment.③Via phagocytosis assay, macrophages and CFSE-labeled ovarian cancer cells were co-cultured with the indicated anti-CD47and phagocytic index was calculated.(4) Analyze the proportion of ALDH+ovarian cancer stem cells by flow cytometry, and explore the expression of CD47in ALDH+/ALDH-ovarian cancer cells.Results (1) CD47highly expresses in human ovarian cancer cell line SKOV-3(CD47mean fluorescence47.44±4.55) and ovarian cancer cells (CD47mean fluorescence17.00±10.93), and they were significantly higher than in normal ovarian tissue cells (CD47mean fluorescence3.03±1.14), and the difference was statistically significant (P<0.05). There is no significant difference in patients age, histological type(P>0.05), and the differences among disease clinical stages and pathological differentiation were significant (P<0.05).②The macrophages induced by human cord blood mononuclear cells than mouse macrophages and human macrophages had complex culture conditions, more variable factors, and phagocytosis in sharp contrast to mouse macrophages and human macrophages were weak. Thus subsequent experiments in vitro were applicated in mouse macrophages and human macrophage cells.(3) Experiments in vitro showed that anti-CD47could significantly promote macrophage phagocytosis of ovarian cancer cells, the difference was statistically significant (P <0.05), and with the antibody itself, and the role of HLA mediated mechanism was no significant relationship (P>0.05). With the change of the concentration of anti-CD47, phagocytic index was a significant concentration dependent manner (P<0.05). When in10μg/mL, the phagocytic was efficiency (phagocytic index of0.35±0.09), which may be related to antibody concentration reached saturation state.(4) Flow cytometry analyzed7cases of ovarian cancer cell specimens, and ALDH+ovarian cancer stem cells was in a very small proportion (1.65±1.27)%. And the CD47expression in ALDH+ovarian cancer stem cells (19.85±3.80) was significantly higher than that of ALDH-ovarian cancer cells (13.06±5.88), and the difference was statistically significant (P<0.05).Conclusion (1) CD47highly expresses in ovarian cancer cells, and it is correlated with the clinical stage of the disease and tumor differentiation.(2) Anti-CD47could significantly promote the phagocytosis in vitro, indicating that it might be a potential target of targeted cancer therapy.(3) ALDH+ovarian cancer stem cells was significantly higher CD47expression than the ALDH-ovarian cancer cells, and provide experimental evidence for research of ovarian cancer stem cells.