Immune Response Of BFGF/VEGF Complex Peptide And Function In Inhibiting The Tumor

Object: Recent research showed that over-expression of VEGF and bFGF intumors plays a key role in tumor angiogenesis and metastasis of cancer cells. In thisstudy, we analyzed the immune response of the VEGF/bFGF complex peptide (VBP3)in immunized C57mice and its function in inhibiting tumors.Methods: VBP3was expressed in E.coli with IPTG inducing and purified withNi-NTA affinity chromatography and anion exchange chromatography. BALB/cmice were immunized with purified VBP3and anti-VEGF; then anti-bFGF antibodieswere prepared. The antibody titer and specificity were analyzed by ELISA. Thepurified antibodies were used to investigate the function of inhibiting blood vesselformation and migration of HUVEC cells; also, the proliferation of SKOV3and LL-2cells were analyzed by CCK-8. The phosphorylation levels of Erk1/2and Akt weredetected by Western blot. C57BL/6mice were only immunized with VBP3and PBS.Following this procedure, LL-2cells were subcutaneously inoculated into the flanksof C57BL/6mice to create tumor graft mice. The Leukocytes were separated from themice tumors and spleen tissue. The expression of CD11b,CD11c,Gr-1were analyzedby flow cytometry. We then researched how VBP3inhibited the growth of tumors.Results: The SDS-PAGE results showed that VBP3protein was expressed at30KDa. ELISA’s results showed that high titer anti-VEGF and anti-bFGF antibodieswere produced in BALB/c mice. These antibodies significantly inhibited the tubeformation and migration of HUVEC cells and the proliferation of SKOV3and LL-2cells. The SKOV3cells’ inhibition rate was about39.20%and the LL-2cells’inhibition rate was about70%at100μg/ml of the antibodies. In addition, theantibodies decreased the phosphralation level of Erk1/2and Akt assayed by theWestern-blot. Compared to the C57BL/6mice immunized with PBS, tumor sizedecreased in the mice immunized with VBP3and the inhibition rate was88.5%. Flowcytometry assays showed that the VBP3vaccination could downregulated themyeloid-derived suppressor cells, which may relate the immune escape mechanism of tumor cells within their bodies.Conclusion: The results revealed that VBP3can inhibit angiogenesis andprevent immune escape. VBP3can also effectively inhibit the proliferation of LL-2cells’ growth in vivo and in vitro and can possibly be used as a vaccine for tumortherapy.