Utilizing VEGF165b mutant as an effective immunization adjunct to augment antitumor immune response.

Compelling evidence has shown that blocking VEGF via monoclonal antibodies may be beneficial in that it not only inhibits tumor angiogenesis but also reduces immune suppression and promotes T cell infiltration into tumors. Herein, we determined whether our recently generated VEGF165b mutant could be used as a co-immunization adjunct to augment the peptide cancer-vaccine- induced immune response in a mouse model of breast cancer. When co-immunized mVEGF165b with the peptide-based cancer vaccine (MUC1, a T-cell epitope dominant peptide vaccine from Mucin1), the VEGF antibody titers increased approximately 600,000-fold in mice. Moreover, the anti-VEGF antibody also reduced the frequency of regulatory T cells (Tregs) in both preventive and therapeutic scenarios. Mechanistically, the decrease of the Tregs population was associated with a remarkably increased MUC-1-specific IFN-γ-producing CD8+ T cells and anti-MUC1 humoral response. Finally, this combination co-immunization produced a superior antitumor response and significantly prolonged survival of tumor-bearing mice. In conclusion, our findings suggest that mVEGF165b may be an ideal immunization adjunct to enhance the immune efficacy of peptide-based tumor vaccines by overcoming immune tolerance.

VEGF165b mutant with a prolonged half-life and enhanced anti-tumor potency in a mouse model.

VEGF165b has been shown to be an effective anti-cancer agent; however, its short half-life limits further application in the clinical field. The development of a mutant VEGF165b with a prolonged half-life is urgently needed for its future application. A mutant VEGF165b was generated by inactivation of its plasmin cleavage site. The mutant and native VEGF165b proteins without purification tags were expressed via the Pichia pastoris expression system followed by purification with a HiTrap heparin affinity chromatography column through optimization of the purification conditions. Furthermore, its binding affinity with VEGF Receptors and its functions in vitro and in vivo were examined. Results showed that the half-life of mutant VEGF165b increased to approximately 10 times (Intravenous), 9.1 times (Intraperitoneal) and 5.4 times (Subcutaneous) greater than that of VEGF165b, and the mutation did not cause significant alteration of VEGFR1 and VEGFR2 binding affinity. Mutant VEGF165b inhibited the proliferation and migration of HUVECs in vitro, similar to the native VEGF165b. In a mouse melanoma model, mutant VEGF165b exhibited stronger anti-tumor activity in comparison with its native counterpart. These results indicate that the mutant VEGF165b had a prolonged half-life and retained the anti-angiogenic activity of the native VEGF165b, suggesting that this novel mutant VEGF165b may be a stronger anti-cancer agent.