Targeting cells that support
tumor growth by administering potent
angiogenesis inhibitors is currently an area of intense interest. In the present study, a unique plasmid vector for the mouse
endostatin gene, pXLG-mEndo, was constructed and evaluated with and without radiation using the
Lewis lung carcinoma (LLC) cell line. The physical properties of the expressed
endostatin protein were validated by PCR, gel electrophoresis, and Western blot.
Enzyme-linked
immunosorbent and immunocytochemical analyses for the therapeutic gene demonstrated that transfected LLC cells secreted the
protein into the medium. Exposure of the cells to 2 gray (Gy) gamma-rays reduced the time to reach the maximum expression level of the
endostatin gene and also increased the amount of secreted
endostatin protein (P<0.001).
Biological activity of the
endostatin was demonstrated by the inhibition of tube formation by human umbilical vein endothelial cells (HUVEC). Based on (3)H-thymidine incorporation,
endostatin expression significantly depressed
DNA synthesis in HUVEC and LLC cells compared to controls transfected with parental vector or no vector (P<0.005). In addition, radiation increased the efficiency of
endostatin-mediated inhibition of both cell types over a 3-day period post-exposure (P<0.05 or less). Intratumoral injection of 100 small mu g pXLG-mEndo combined with 10 Gy radiation significantly delayed LLC
tumor growth, especially when each modality was delivered twice (P<0.05 or less compared to all other groups). No toxicity was observed. These findings are very promising and suggest that
endostatin therapy with a plasmid vector, such as pXLG-mEndo, may enhance the efficacy of
radiotherapy for
lung cancer.