Experimental Study Of Dasatinib And DC Vaccine In The Treatment Of Breast Cancer In Mice

Objective:To investigate the combined anti-tumor effects of DC vaccine and dasatinib on the treatment of breast cancer in BALB/c mice bearing 4T1 breast cancer cells.On one side,DC vaccine can directly enhance the immune response of anti-tumor.On the other side,dasatinib was proved to be the ability of recruiting T cell into tumor microenvironment,which can indirectly led to decrease the proliferation and metastasis of tumor cells.This investigation provide a theoretical basis for dasatinib and DC vaccine in the treatment of breast cancer.Methods:1.The DC precursors were isolated from both leg bones in mice.Then the cells were cultured with mouse-derived GM-CSF and mouse-derived IL-4,and a large number of bone marrow derived DCs were harvested after bearing with 4T1-lysated liquid and matured by LPS.The bio-immunological features and functions of DC in different stages were investigated by using methods of inverted microscope and flow cytometry.2.To investigate the adequate number of 4T1 cells,and then establish breast cancer mice model.To investigate the adequate dose of dasatinib in the capacity of enhancement of CD8+T cells into tumor microenvironment.DC vaccine by peritumoral injection or/and dasatinib by oral were applied for the treatment of breast cancer mice model.On 34 days after 4T1 cells injected,in order to assess the effect of different treatments on mice,tumor size,lung metastasis,and animal survival were monitored.Ki-67,CD8,and NKG2 D expression difference in tumor cells were evaluated by immunohistochemical analysis and the expression difference of apoptosis were evaluated by Tunel stain method.Based on the methods mentioned above and results,assess there's possible combined antitumor effect of dasatinib and DC vaccine on the treatment of metastatic breast cancer and explore the mechanism.Results:1.The expression of CD11 c,CD86,MHCII?CD80 of Mature DCs using flow cytometry showed(92.3±1.2)%?(86.4%±2.3)?(87.1±1.8)%?(89.7±2.5)%?The expression of CD8 in 0mg/kg,5mg/kg,10mg/kg,15mg/kg,30mg/kg dasatinib mice using immunohistochemical method showed(17.80±1.09)?(18.01±3.89)?(19.78±2.49)?(46.32±4.04)?(47.75±3.56)?2.The size of tumor in DC vaccine group,dasatinib group were reduced compared to control group,but there's no significantly difference(P >0.05).Compared combined group to dasatinib group,DC vaccine or control group,the size of tumor was significantly reduced(P<0.05).The mass of spleen in combined group was the lightest,whereas the mass of spleen in control group was the heaviest,and there's significantly difference(P<0.05).The mass of spleen between dasatinib group,DC vaccine group and control group showed no significantly difference(P>0.05)?3.Mice treated with DC vaccine or dasatinib showed the longer survival days than mice treated with control group,but there's no significantly differences(P>0.05)?Mice treated with combined treatment compared to other mice showed no statistically significant(P>0.05).4.The results of immunohistochemical method: the expression of Ki-67 and CD31 in combined group were siginificantly decreased than that in DC vaccine,dasatinib,or control group(P<0.05).The expression of Ki-67 and CD31 between dasatinibatinb and DC vaccine were slightly decreased than that in control group,but there's no significantly difference(P<0.05).The number of CD8+ and NK2 GD in primary tumor in combined group were significantly higher than that in other three group,especially the number of CD8+T cells(P<0.05).5.The number of the lung metastatic cancer nodules were counted by dissecting microscope showed to be significantly lower in dasatinib plus DC vaccine treated mice than in dasatinib alone,DC vaccine alone and the untreated control mice,and there's significantly difference(P<0.05).Microscopic analysis of TUNEL assay exhibited a significant increase of apoptotic cell in mice treated with the combined treatment than any other group(P<0.05).The difference among DC vaccine group,dasatinib group and control group exhibited no statistically significant(P>0.05).Conclusions:The combined therapy using DC vaccine and dasatinib in treatment of metastatic breast cancer showed enhancement of the activity of immune cells,inhibitition the growth of tumor cells and reduction of the number of lung metastasis,but not the prolong the survival of the tumor-bearing mice.In this study,the combination drug therapy does produce a synergistic effect and its antitumour effect may be caused by the enhancement of immune cells(CD8+T and NK cells)into tumour microenvironment.The study provided the experimental basis for the treatment of metastatic breast cancer in the further.