Synergistic anti-tumor effect of recombinant chicken fibroblast growth factor receptor-1-mediated anti-angiogenesis and low-dose gemcitabine in a mouse colon adenocarcinoma model

Abstract

Aim: To evaluate whether the combination of recombinant chicken fibroblast growth factor receptor-1 (FGFR-1) protein vaccine (cFR-1) combined with low-dose gemcitabine would improve anti-tumor efficacy in a mouse CT26 colon adenocarcinoma (CT26) model.

Methods: The CT26 model was established in BABL/c mice. Seven days after tumor cell injection, mice were randomly divided into four groups: combination therapy, cFR-1 alone, gemcitabine alone, and normal saline groups. Tumor growth, survival rate of tumor-bearing mice, and systemic toxicity were observed. The presence of anti-tumor auto-antibodies was detected by Western blot analysis and enzyme-linked immunospot assay, microvessel density (MVD) of the tumors and tumor cell proliferation were detected by Immunohistochemistry staining, and tumor cell apoptosis was detected by TdT-mediated biotinylated-dUTP nick end label staining.

Results: The combination therapy results in apparent decreases in tumor volume, microvessel density and tumor cell proliferation, and an increase in apoptosis without obvious side-effects as compared with either therapy alone or normal control groups. Also, both auto-antibodies and the antibody-producing B cells against mouse FGFR-1 were detected in mice immunized with cFR-1 vaccine alone or with combination therapy, but not in non-immunized mice. In addition, the deposition of auto-antibodies on endothelial cells from mice immunized with cFR-1 was observed by immunofluorescent stain-ing, but not on endothelial cells from control groups. Synergistic indexes of tumor volume, MVD, cell apoptosis and proliferation in the combination therapy group were 1.71 vs 1.15 vs 1.11 and 1.04, respectively, 31 d after tumor cell injection.

Conclusion: The combination of cFR-1-mediated anti-angiogenesis and low-dose gemcitabine synergistically enhances the anti-tumor activity without overt toxicity in mice.

Figure 1

The combination therapy inhibited tumor growth (A) and improved the survival of tumor-bearing mice (B) significantly. ^aSignificant difference compared to untreated group, ^aP < 0.05.

Figure 2

Induction of auto-antibodies by cFR-1 vaccine alone or combination therapy as assessed by Western blot analysis.

Figure 3

The deposition of auto-antibodies on the endothelial cells by immunofluorescent staining (A-B) and the numbers of APBCs in spleens of mice by ELISPOT assay (C) in cFR-1 vaccine alone or combination therapy.

Figure 4

The combination therapy synergistic inhibition of angiogenesis (A) and proliferation (B), and induction apoptosis (C) at d 31 after tumor cell injection. ¹Synergistic relationship in the combination therapy (synergistic index > 1). ^aP < 0.05 vs untreated group.

Figure 5

HE staining of heart (A), liver (B), kidney (C) and lung (D) in recipient mice. No organic hemorrhage appeared in the combination therapy group and no differences were found among of C + G (a), cFR (b), G (c), and NS groups (d).