[Anti-metastatic effect of vascular endothelial growth factor receptor 2 extracellular domain gene-modified dendritic cell vaccination in murine model with experimental pulmonary metastasis].
OBJECTIVE
To investigate the anti-metastatic effect of vascular endothelial growth factor receptor 2 extracellular domain gene-modified dendritic cell (DC-sVEGFR-2) vaccination.


METHODS
Dendritic cells (DC) were electroporated with pcDNA3. 1/sVEGFR-2 plasmid DNA. Expression of sVEGFR-2 was determined by ELISA. For immunization, C57BL/6 mice were intravenously injected three times with 1 x 10(5) cells per mouse of DC, pcDNA3. 1-transfected DC (DC-vector) , DC-sVEGFR-2, or 100 microl of PBS at 7-day intervals. At 10 days after the last immunization, the immunized mice were subjected to assessment of cytotoxic T lymphocyte ( CTL) response to VEGFR-2, alginate bead analysis of tumor cell-induced angiogenesis, and observation of the anti-metastatic effect in B16 melanoma metastasis model. CTL activity was determined by a standard 4-h 51Cr release assay against VEGFR-2 + vascular endothelial cell line H5V, 3LL cells stably transfected with pcDNA3. 1/sVEGFR-2 (3LL,-sVEGFR-2), and VEGFR-2- cell lines EL-4 and 3LL. Monoclonal antibodies GK1.5 anti-CD4 and 2.43 anti-CD8 were used to deplete in vivo CD4 + T cells and CD8' T cells, respectively.


RESULTS
DC-sVEGFR-2 could effectively express sVEGFR-2, whereas DC-vector and DC could not. Immunization of mice with DC-sVEGFR-2 significantly induce CTL activity against VEGFR-2 + cell lines H5V and 3LL-sVEGFR-2, however, no significant CTL activity was observed when VEGFR-2- syngeneic cell lines EL-4 and 3LL. were used as target cells, implying this CTL activity was VEGFR-2 specific. Alginate bead analysis of in vivo neoangiogenesis showed that the inhibition reached 50% in mice vaccinated with DC-sVEGFR-2 compared with mice vaccinated with DC, DC-vector or PBS. Anti-metastatic experiment showed that profound reduction in pulmonary metastases was found in mice immunized with DC-sVEGFR-2, while mice immunized with PBS, DC, DC-vector developed extensive pulmonary metastases. The number of tumor nodules on lung surface decreased by 81.9% in mice immunized with DC-sVEGFR-2 when compared with mice immunized with DC-vector (49.7+/-12.7 vs. 9.0+/-3.2). In vivo T cell subset depletion experiments showed that the anti-metastatic effect of DC-sVEGFR-2 vaccination was abrogated in CD8 + T cell-depleted but not in CD4+ T cell-depleted mice.


CONCLUSIONS
Immunization of mice with DC-sVEGFR-2 could break self-tolerance and induce a significant CTL response to VEGFR-2, leading to profound inhibition of tumor-cell induced angiogenesis and metastasis. This anti-metastatic effect is mainly mediated by CD8+ T cells.
Pan
Jian-ping
JP
Institute of Immunology, Zhejiang University, Hangzhou 310031, China. jppan@zju.edu.cn
Weng
Yue-song
YS
Wu
Qian-qian
QQ
D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D019496:Cancer Vaccines / Q000276:immunology*; D018827:Carcinoma, Lewis Lung / Q000098:blood supply / Q000276:immunology / Q000473:pathology; D045744:Cell Line, Tumor; D003713:Dendritic Cells / Q000276:immunology* / Q000378:metabolism; D018274:Electroporation; D005260:Female; D016219:Immunotherapy, Adoptive / Q000379:methods*; D008175:Lung Neoplasms / Q000556:secondary / Q000628:therapy*; D051379:Mice; D008810:Mice, Inbred C57BL; D009389:Neovascularization, Pathologic / Q000276:immunology / Q000628:therapy*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000096:biosynthesis / Q000235:genetics*
D019496:Cancer Vaccines; D040301:Vascular Endothelial Growth Factor Receptor-2