Tumor Microvessel Endothelial Cells As A Target For Gene Therapy Of Cancer And Microcirculation Experiments

Tumors require ongoing angiogenesis to support their growth, and inhibition of angiogenesis has been recognized as a viable approach for anticancer therapy. Nowdays, antiangiogenic tumor therapy has attracted intense interest. To promote tumor regression and maintain dormancy, antiangiogenic agents in protein therapy should be chronically administered. Gene therapy offers a potential way to achieve sustained therapeutic release of potent angiostatic factors, and has achieved the same goal like protein therapy with lower expense. Of so many antiangiogenic strategies, most target the cancer cells, lacking the study of tumor neovascular network and microcirculation. As a step toward serial studies of mouse Endostatin (ES) gene therapy on tumor growth, metastasis and tumor microcirculation, we carried out the following experiments:1. We cloned the Endostatin cDNA by RT-PCR from Chinese Kunming mouse liver RNA and varified it by sequencing, constructed pSecTag2-ES (ES) and pSecTag2-ES-TT (ES-TT, TT derived from tumor neovasculature-targeting peptide) eukaryotic expression plasmids, transfected them into COS-7 cells to achieve fusion protein expression. The supernatants of such transfected cells induced the apoptosis and inhibited the proliferation of endothelial cells in vitro. The biological activity of the expressed ES-TT on the proliferation of endothelial cells was lower than that of the expressed ES (p<0.05).2. A modified chorioallantoic membrane assay of 9-day chick embryos was employed to evaluate the anti-angiogenesis effect of ES gene transfer, and the suppressed formation of regional microvascular network was observed after ES gene transfer onto the chorioallantoic membrane for a continuing hatch of 3 days.3. To assess the effects of ES gene therapy on tumor growth and metastasis, plasmid DNA and DNA/PVP40 complex were administered by intramuscular injection to C57 mice bearing primary subcutaneous Lewis lung carcinoma. The expression of ES and ES-TT fusion protein in serum was detected by ELISA, while there existed no statistic difference of fusion protein within the tumor extracts between ES/PVP40 and ES-TT/PVP40 group. The electron transmission...