An enhanced immune response against G250, induced by a heterologous DNA prime‑protein boost vaccination, using polyethyleneimine as a DNA vaccine adjuvant.

The heterologous DNA prime‑protein boost vaccination approach has been widely applied as an immune treatment for carcinoma. However, inefficient delivery of DNA remains a major issue. In the present study, polyethyleneimine (PEI) was used as a DNA vector carrier to improve the transfection efficiency of the DNA vaccine and stimulate the humoral and cellular immunity against the renal carcinoma‑associated antigen G250. A protein vaccine was included in the immunization strategy in order to produce a prime‑boost effect. A DNA plasmid encoding an antigen G250 fragment was constructed and complexed with PEI. A protein vaccine against G250 was expressed in BL21 (DE3) Escherichia coli cells, by transformation with a pET28a(+)/C‑G250 plasmid. The protein was purified using a nickel‑nitriloacetic acid purification system. The in vitro transfection efficiency of the DNA vaccine was analyzed in HEK293 human endothelial kidney cells. The in vitro transfection efficiency in HEK293 cells was highest 48 h after transfection. Furthermore, mice were primed with 200 µg pVAX1/C‑250 plasmid complexed with PEI, and boosted using 50 µg of purified C‑G250 protein. In order to evaluate the immune response the antibody titer, splenocyte response, and interferon‑γ levels from CD8+ T‑cell splenocytes were analyzed using ELISA, lymphocyte proliferation or enzyme‑linked immunosorbent spot assays. Firstly, the pVAX1/C‑G250 plasmid was shown to be constructed successfully. As compared with the DNA group, the antibody titer, lymphocyte proliferation percentage, and cytokine production level induced by the DNA‑PEI and DNA‑PEI+C‑G250 groups were significantly higher. Furthermore, the DNA‑PEI+C‑G250 group exhibited the strongest humoral and cellular response. Owing to the adjuvant effect of PEI, the pVAX1/C‑G250‑PEI prime plus C‑G250 protein boost regimen could induce a strong immune response, and has been proved to be a potent vaccine candidate against renal cell carcinoma.