DC/melanoma Fusion Cells And Rapamycin Stimulated CTLs Specificallyagainst Melanoma Research

Background Currently the four means of treating cancer are surgery,radiotherapy,chemotherapy and biological therapy.Tumor immunotherapy has become a hotspot of biological treatment.Dendritic cells(DCs)that are the body’s most effectively professional antigen-presenting cells have important value in tumor immunotherapy.In recent years,researchers have made some attempts to use different forms of tumor antigens such as peptides,tumor cell lysates,tumor cell apoptosis products of genetic engineering or tumor antigen genes into dendritic cells via a vector to sensitize DCs.Sensitized DC vaccines in vivo have induced specific cytotoxic T cells(CTLs)generation,effectively stimulated the body’s specific anti-tumor immunity,and achieved certain effect in the treatment of malignant tentative.However,little is known about tumor antigens,so researchers have used polyethylene glycol method(PEG)method to fuse directly DCs and tumor cells,informing DC/tumor fusion cells.The fusion cells in the induction of anti-tumor immune response have its unique advantages,which can express all known and unknown tumor antigens,allowing activation of T cells in vitro and promoting generation of tumor antigen-specific cytotoxic T lymphocytes.DC/tumor fusion cells can express both tumor antigens and DC surface molecules,which can more effectively stimulate T cell activation,proliferation and differentiation.Related research reports have indicated that rapamycin(Rapamycin,Rapa)can also exhibits immunostimulatory effects on memory CD8~+T cell differentiation.The drug improved both quantity and quality of memory CD8~+T cells.At present,there is no literature report that DC/B16 fusion cells combining with rapamycin co-induce T cells can stimulate the generation of a large number of long-lasting effective and survival ability of tumor antigen-specific cytotoxic T lymphocytes,then effectively and specifically kill tumor target cells.Objective To explore the proliferation and the ability to kill target cells of tumor antigen-specific cytotoxic T lymphocytes induced by DC/B16 fusion cells and rapamycin and furtherly verify that its mechanism of tumor antigen-specific CTL in vivo against melanoma.Methods Firstly DC/B16 fusion cells were prepared.Tumor antigen-specific CTLs were induced by DC/B16 fusion cells and rapamycin(referred to as DC/B16-Rapa-T cells),whose proliferation and ability to kill target cells were analyzed by flow cytometry.In addition,ELISPOT measured the positive cells secreting IFN-γcytokine.Finally melanoma mice model was constructed and adopted DC/B16-Rapa-T cells to tumor-bearing mice and observed anti-tumor effects of adoptive therapy and explore furtherly its mechanism.Results T cell proliferation assay showed that comparing specific T cells alonely induced by DC/B16 fusion cells(DC/B16-T cells)and specific T cells alonely induced by rapamycin(Rapa-T cells)with tumor antigen-specific CTLs induced by DC/B16 fusion cells and rapamycin(DC/B16-Rapa-T cells),DC/B16-Rapa-T cells were the most effective in promoting T cell activation and proliferation.ELISPOT assay showed that positive cells secreting IFN-γcytokines of DC/B16-Rapa-T cells were more than T cells alone,Rapa-T cells and DC/B16-T cells.Cytotoxic assay indicated that DC/B16-Rapa-T cells could kill B16 cells effectively and specifically.DC/B16-Rapa-T cells were adopted to mice bearing melanoma,which could effectively inhibit the proliferation of melanoma cells and prolong the survival time of mice bearing melanoma.Compared with the other groups,DC/B16-T cells could decrease the amount of CD4~+CD25~+CD127~-regulatory T cells(Tregs)and Myeloid-derived suppressor cells(MDSCs).Conclusion Tumor antigen-specific CTLs induced by DC/B16 fusion cells and rapamycin could efficientiy,durablely and specifically kill tumor target cells.Therefore,this novel adoptive immunotherapy method could improve the anti-tumor effects of T cells.