Alpha-galactosylceramide loaded tumor cells in combination with TLR9 agonists induce potent anti-tumor responses in a murine model of colon cancer

Abstract

Objective To design a new cancer vaccine by using alpha-galactosylceramide （α-Galcer,α-GC） loaded tumor cells in combination with TLR 9 ligand and to evaluate its therapeutic effects on colon canc-er in mice.Methods MC38 cells were transfected with lentivirus （GFP-CD1d） to prepare CD1d-MC38 cells. The expression of CD1d molecules in CD1d-MC38 cells was detected by fluorescence microscopy , RT-PCR and flow cytometry.The sorted CD1d-MC38 cells were loaded with α-Galcer to prepare CD1d-MC38/α-GC complex. Flow cytometry was performed to evaluate the efficiency of combination .A mouse model of colon cancer was es-tablished to investigate the therapeutic effects of α-Galcer loaded tumor cells in combination with TLR 9 ligand （ CD1d-MC38/α-GC＋CpG1826） on colon cancer in mice by analyzing tumor growth and mice survival time .Im-munohistochemical staining was used to detect CD 4＋T and CD8＋T infiltrating lymphocytes in tumor tissues .Re-sults The MC38 cancer cells that expressed CD 1d and GFP were successfully constructed , among which 98.10%±2.53%were positive for CD1d.Moreover, the CD1d-MC38 cells could combine with α-Galcer effec-tively in a dose and time dependent manner .Compared with PBS treated group ,α-GC treated group and TLR9 ligand treated group , the experimental vaccine strategy was sufficient to inhibit the growth of established tumors and prolong survival of tumor-bearing mice （P〈0.01）.Immunohistochemistry analysis revealed that levels of CD4＋T cells and CD8＋T cells in experiment group were significantly higher than those in groups treated with PBS,α-GC and TLR9 ligand （P〈0.01）.Conclusion CD1d-MC38/α-GC in combination with CpG1826 could efficiently inhibit the growth of established tumors and prolong survival of tumor-bearing mice .Immunohisto-chemistry analysis revealed that CD 4＋T cells and CD8＋T cells played important roles in anti-tumor immunity. Key words: iNKT cells; Dendritic cells; α-Galcer; Cancer vaccine