| Objective:To revealed the immunotherapeutic effect and mechanism of CpG modified with mannose and lipid.Methods:CpG were modified with lipid and mannose by chemical synthesis method,and the novel adjuvant was termed as LM-CpG@Au.OVA protein was used as model antigen and melanoma model was established to test the immunotherapy effect of the adjuvant.In tumor model,the antitumor effect and mechanism of LM-CpG@Au on the response of CTLs were examined by flow cytometry and cell cytotoxicity assay.The effects of LM-CpG@Au on macrophage polarization and Tregs differentiation in tumor microenvironment were also studied by cell depletion assay and cytokine neutralization assay.We also tested the therapeutic effect of the combination of the adjuvant and anti-PD-1 treatment.Results:Tumor growth was significantly inhibited by LM-CpG@Au.The percentages,total numbers and tumor-specific killing capacity of CTLs in spleen were also enhanced by LM-CpG@Au treatment.The percentages of CTLs in tumor microenvironment and the numbers of infiltrating lymphocytes per gram of tumor tissue in LM-CpG@Au treatment group were significantly higher than other groups.The secretion of IFN-γ and TNF-α by CTLs in the spleen and tumors from LM-CpG@Au treated group were also significantly enhanced.Cell depletion assay and cytokine neutralization assay revealed that the antitumor effect of LM-CpG@Au was depended on IFN-γ derived from CTLs.In the tumor microenvironment,the percentage of M1 macrophage was significantly increased after LM-CpG@Au treatment,while the percentage of M2 macrophage and Tregs were significantly reduced.Depleting CTLs or neutralizing IFN-γ abrogated the effects of LM-CpG@Au on M1 polarization and Treg regulation.Co-culture experiment of T cells and tumor tissues also showed that LM-CpG@Au regulated the polarization of macrophages and the differentiation of Tregs by CTLs.Finally,we found that anti-PD-1 therapy could further enhance the therapeutic effects of LM-CpG@Au.Conclusion:LM-CpG@Au can significantly inhibit the growth of tumors.LM-CpG@Au activated tumor-specific CTLs,promoted the migration of CTLs to tumor,and regulated the tumor microenvironment through CTLs and cytokines secreted by CTLs. |
