Dendritic Cells Modified By Recombinant Adeno-associated Virus Encoding Extracellular Fragment Of Epidermal Growth Factor Receptor Variant â…¢ Induce Antigen-specific CTL Response To EGFRvâ…¢-expressing Glioma Cells

Background: Conventional immunotherapies for malignancy can’t succeed in targeting tumor cells exclusively so that their efficacy ultimately limited by nonspecific toxicity. Immunologic targeting of tumor-specific antigens of tumor cell surfaces may allow more precise eradication of neoplastic cells. The epidermal growth factor receptor variant III (EGFRvⅢ) is an attractive therapeutic target as it frequently expresses in glioma and many other types of cancers.Objective: To investigate the selective killing effect of T lymphocytes induced by Epidermal Growth Factor Receptor Variant Ⅲ (EGFRvⅢ)-specific dendritic cells against EGFRvⅢ+glioma U87cells in vitro.Methods: Our current study aimed that we used the rAAV system to modify dendritic cells (DCs) with the extracelullar fragment of epidermal growth factor receptor variant III (EGFRvⅢext) to induse specific CTLs against EGFRvⅢ. Then we investigated the effect of the CTLs in killing the Glioma cells expressing EGFRvⅢ in vitro: the expression of CD80、CD83and HLA-DR on DCs by flow cytometry, the expression of EGFRvIIIext protein in DCs by western blot, the selective killing by MLR and the serection of IL-2and IFN-γ.Results: The results show that (1) The expressions of CD80, CD86and HLA-DR on DCs were81.59%,67.37%and85.38%analyzed by flow cytometry.(2) The transduction efficiency of rAAV/EGFRvⅢext/GFP on DCs reached around91.37%and expression of EGFRvⅢext protein was verified by immunoblotting as a band of about58kDa.(3) MLR demonstrated specific and efficient cytotoxicity of EGFRvⅢext+DCs indused CTLs against EGFRvⅢ expressing U87cells.(4)A robust increase in the IFN-γ and IL-2secretion was detected in the co-culture supernatant of the EGFRvⅢext+DCs indused T cells and the EGFRvⅢ expressing U87cells.(5) High proliferation of T cell populations. Conclusion: Our study demonstrates that EGFRvⅢext gene-transduced DCs can efficiently enhance immunity against gliomas expressing EGFRvⅢ in vitro.