| Objective: Inhibition of angiogenesis has been shown to be an effective strategy in cancer clinical therapy now.MichaelS.O'Reilly has found endostatin in the urine of tumor-bearing mice in 1997. It is reported that endostatin could suppress tumor growth effectively by inhibiting of angiogenesis targeted different pathways.Howerer,its widespread application has been hampered by difficulties in the large-scale production of the antiangiogenic protein,rapid loss bioactivity of the protein,the cumbersome dairy administration. These difficulties could be overcome through transfer of the endostatin gene. It has been the hotspot to expressing endostatin mediated by adenviral vecto. We have constructed a recombinant adenovirus that express human endostatin. We introduce a new therapy strategy in which recombinant adenovirus are first used to infect fibroblasts to make them to secrete endostatin,which could inhibit the viability of ECV304 cells in vitro. Then the infected fibroblasts are used to administrate the tumor-bearing mice to inhibit the growth of tumor by the secreted endostatin. The aim of this study was to detect the concentration of endostatin secreted by the fibroblasts infected with recombinant adenovirus and to investigate the effect of the infected fibroblasts combined with cisplatin(DDP) on the growth of xenografted malignant melanoma in nude mouse.Methods: (1) The concentration of endostatin ,secreted by the fibroblasts infected with recombinant adenovirus, was detected in vitro and in vivo by enzyme linked immunosorbent assay(ELISA). (2) After the establishment of the model, nude mouse were divided into 7 groups(five in each) receiving respectively intraperitoneally or subcutaneouly for 10 to 14 days: fibroblasts infected with recombinant adenovirus carrying human endostatin gene, recombinant adenovirus carrying human endostatin gene ,cisplatin(DDP), infected fibroblasts combined with DDP, recombinant human endostatin(YH-16, endostar), recombinant human endostatin combined with DDP,and fibroblasts infected with adenovirus carrying none gene. On 21th day tumor volume was measured. The tumor inhibition rate(TIR) was calculated. The expression of VEGF and microvessels(MVD) was examined by immunohistochemistry.Result: (1) 24 hours after systemic administration of the fibroblasts infected with recombinant adenovirus to mice,the serum concentration of endostatin was 113.4ng/ml,then remained above the level before administration until the sixth day. (2)TIR of the infected fibroblasts group, the recombinant adenovirus group, the DDP group, the YH-16 group, the fibroblasts combined with DDP group and the YH-16 combined with DDP group was respectively 53.41%,35.06%,45.72%,17.52%,84.64% and 59.40%. Infected fibroblasts significantly reducted the growth rates of tumors compared with the recombinant adenovirus group and recombinant human endostatin (p<0.05) . DDP combined with the infected fibroblasts or YH-16 significantly reducted the growth rates of tumors compared with the infected fibroblasts only(p<0.05) , YH-16 only(p<0.05) or DDP only(p<0.05). Immunohistochemistry staining of the tumors demonstrated a decreased number of blood vessels and express of VEGF in the treatment groups versus the controls(p<0.05). The MVD in the infected fibroblasts group was decreased compared with the recombinant adenovirus group and recombinant human endostatin (p<0.05) . DDP combined with the infected fibroblasts significantly reducted the MVD and express of VEGF compared with the infected fibroblasts only(p<0.05) or DDP only(p<0.05).Conclusion: Persistent effective serum concentrations of endostatin are achived after systemic administration of the fibroblasts infected with recombinant adenovirus carrying human endostatin gene to mice. Infected fibroblasts has more effective inhibitory activity against the growing of malignant melanoma than recombinant adenovirus and recombinant human endostatin ,and has synergistic effect in the combination with DDP. |
