Anti-tumor Effect Of Dendritic Cell Vaccines Transfected With Total RNA Of Lung Adenocarcinoma Cell And Induced By Modified Cytokine Cocktail In Vivo

Objectives: In view of the current status that dendritic cell vaccine can’t effective induce antitumor immune response, we modifies the conventional cytokine cocktail by utilizing Toll-like receptor(TLR) ligands in order to prepare a more effective DC vaccines for clinical application. By establishment of the tumor-bearing nude mice model, we explore the specific antitumor effect of DC vaccine induced by different cytokine cocktails on transplanted tumor.Methods: Peripheral blood mononuclear cells(PBMC) were isolated from peripheral blood stem cells and induced into immature DC. DC were transfected with total RNA of human lung adenocarcinoma cell line A549 through electroporation, then stimulated with conventional cytokine cocktail(IL-6、IL-1β、TNF-α、PGE2) and modifiedcytokine cocktail( IL-1 β、 IL-6、 TNF-α、PGE2、 poly I:C、 Cp G ODN), respectively. Antigen-sensitized DC were cocultured with autologous T cells to activate antigen specific cytotoxic T lymphocyte(CTL). The phenotypes of T cells were analyzed by flow cytometry, the secretion of IFN-γ、TNF-α、IL-4、IL-5 and IL-10 from T cells were measured by enzyme-linked immunosorbent assays(ELISA). After establishment of the A549 implanted lung adenocarcinoma model, nude mice were randomly divided into empty control group, immature group, conventional group and modified group. Peritumoral injection of CTL was performed on day 15, day 22 and day 29 after inoculation of tumor cells. The volumes and weights of tumors were measured. The expression of COX-2、VEGF、Bcl-2 and Bax in the tumor tissues were determined using immunohistochemistry and Western blotting.Results:(1) The percentage of CD3+CD8+ T cells in modified group was raised significantly with(63.98 ± 1.96) %, higher than those of immature group with(35.00 + 3.24) % and conventional group with(57.10 + 2.69) %(P <0.05).(2) T cells in modified group produced much higher levels of Th1 cytokines IFN-γ and TNF-α than immature group and conventional group(P <0.05), Th2 cytokines IL-10 in modified group and conventional group are slightly higher than immature group(P<0.05), nevertheless there was no significant difference between two groups(P >0.05); IL-4 and IL-5 in each group is undetectable.(3) After immunotherapy with CTL, tumor size of nude mice in modified group was(203 ± 52) mm3 and was significantly inhibited compared with the immature group(512 ± 33) mm3 and the conventional group(345 ± 63) mm3(P <0.05). Moreover, the inhibitive ratio of tumor weight in modified group was the highest with 69.62 %.(4) The expression of Bax in modified group was significantly up-regulated and the expressions of COX-2, VEGF and Bcl-2 were down-regulated.Conclusion: DC vaccines which are electroporated with total RNA of lung adenocarcinoma cell and induced by modified cytokine cocktail can effectively inhibit the growth of lung adenocarcinoma transplanted tumors in nude mice through significant induction type Th1 immune response and production effective antigen specific CTL.