Construction of a DNA vaccine encoding Flk-1 extracellular domain and C3d fusion gene and investigation of its suppressing effect on tumor growth.
Although the critical role of complement component C3d as a molecular adjuvant in preventing virus infection is well established, its role in cancer prophylaxis and treatment is unclear. In this study, we constructed a recombinant plasmid encoding Flk-1 and C3d3 fusion proteins and investigated its transient expression in vitro in transfected eukaryotic cells and its antibody response in immunized mice. Subsequently, we investigated the vaccine's ability to elicit an immune response leading to suppression of angiogenesis and tumor growth in mice bearing bladder transitional cell carcinoma. Using Western blotting, immunocytochemistry, and flow cytometry, we detected the expression of Flk-1 and C3d3 fusion proteins in COS-7 cells transfected with these recombinant plasmids. Further binding experiment using CR2 (C3d receptor) positive Raji cells that were incubated with transfected COS-7 supernatant indicated that C3d was successfully fused to Flk-1. Although both vaccines elicited peak antibody levels at 5 weeks, Flk-1-specific antibody titer in pSG.SS.Flk-1(ECD).C3d3.YL-immunized mice was significantly higher when compared to pSG.SS.Flk-1(ECD).YL-immunized mice. The results of experiments with bladder tumor-bearing mice showed that the vaccine inhibited tumor growth significantly. These results suggest that C3d plays a critical role in tumor immunotherapy by promoting antibody response in Flk-1-based DNA vaccines. This approach may provide a new strategy for the rational design of anti-angiogenic therapies for the treatment of solid tumors and provide a basis for the further exploitation and application of the anti-angiogenesis DNA vaccines.
Liang
Pei-he
PH
Department of Urology, Daping Hospital, The Third Military Medical University, 10 Changjiang Branch Road, Yuzhong District, Chongqing, China. lph1972@163.com
Zhang
Ke-qin
KQ
Xu
Gui-lian
GL
Li
Yan-feng
YF
Wang
Luo-fu
LF
Nie
Zhi-lin
ZL
Ye
Jin
J
Wu
Gang
G
Ge
Cheng-guo
CG
Jin
Feng-shuo
FS
D000276:Adjuvants, Immunologic; D000818:Animals; D019496:Cancer Vaccines / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use; D002295:Carcinoma, Transitional Cell / Q000276:immunology / Q000473:pathology / Q000628:therapy; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D015933:Complement C3d / Q000235:genetics / Q000276:immunology*; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D009368:Neoplasm Transplantation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D001749:Urinary Bladder Neoplasms / Q000276:immunology / Q000473:pathology / Q000628:therapy; D019444:Vaccines, DNA / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics / Q000276:immunology*
D000276:Adjuvants, Immunologic; D019496:Cancer Vaccines; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA; D015933:Complement C3d; D040301:Vascular Endothelial Growth Factor Receptor-2