| T cell mediated immunity plays a key role in antitumor responses, and it is also the major target of antitumor immunotherapy. CTLA-4, a second T cell receptor for B7, which is expressed on the surface of activated T cells, has been shown to play an inhibitory role in regulation of T cell responses by preventing full T cell activation or terminating T cell responses. Also, it has been shown that antibody-mediated CTLA-4 blockade can augment host antitumor responses by prolonging T cell activity and/or facilitating antigen-specific T cell costimulatory activation. Another approach taken to raise antitumor immune responses has been to use cytokine-expressing tumor cells as vaccines, which in turn have paracrine effects on T cells or APCs. F1t3 Ligand(FL), a newly found cytokine, which is one of the most important Dendritic Cells (DCs) growth factors, can expand and activate DCs and some hemaprogenitors in vitro and in vivo. While DCs, the most important APCs, are instrumental for the activation of tumor antigen-specific immune response. Recently, it has been reported that FL can promote antitumor activity through inducing a profound expansion of antitumor effect cells (DCs, NKs, CTLs). However, it has been shown that these treatment regimens alone cannot induce complete tumor regression in wealdy immunogenic tumors. And the exact mechanism(s) involved in the antitumor responses elicited by CTLA-4 blockade remains to be elucidated, there also exists different opinions about it. In the present study, we test the hypothesis that FL- and B7-transduced tumor vaccine therapy, combining with CTLA-4 blockade, should enhance antitumor responses in the treatment of poorly immunogenic tumor. We argue the exact mechanism(s) involved in the antitumor responses elicited by anti-CTLA-4; try to open up a novel avenue for the treatment of weakly immunogenic tumors. We constructed a plasmid pGFP-FL, containing eukaryotic expressing GFP and FL gene, and established a Hepal-6 murine hepatoma cell line (H2) harboring the plasmid, which can express both GFP and FL. The H2 cells were further used as FL-expressing tumor vaccines. We also prepared purified anti-muCTLA4 antibodies using a commercial hybridoma cell line, which can secrete anti-muCTLA4. Simultaneously, based on our former works, we produced an anti-huCTLA4-secreting 7H9B5A8 hybridoma cell line using the classical protocols for producing monoclonal antibodies. This cell line may be beneficial to the future researches about humanizing murine anti-huCTLA4 for clinical antitumor immunotherapy. In the treatment of Hepal-6 bearing C57BL/J mice, we found that, FL- and B7-expressing tumor vaccine, wide type Hepal-6 vaccine, or anti-muCTLA-4 alone all showed inefficient effect against Hepal-6 tumors. And Hepal-6 tumors can be fully rejected using both anti-CTLA4 and FL-expressing tumor vaccines, while no synergizing effects were found between anti-CTLA4 and Hepal-6 vaccine, or FL- and B7-expressing tumor vaccines. Notably, we also found that anti-CTLA4 can well synergize with B7-expressing tumor vaccines. This result is different with that of Hurwitz et at obtained from a similar research in 1998, in which they found that anti-CTLA4 can not synergize with B7-expressing tumor vaccines. And, all mice that displayed complete tumor rejection in different treatment groups were immune to rechallenge of 10-fold Hepal-6 cells when take 30 days post tumor regression. Taken together the results of others and our relevant research, we propose that, anti-CTLA4 can simultaneously e...
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