Identification of novel targets for antiangiogenic therapy by comparing the gene expressions of tumor and normal endothelial cells.
Bottom Line: Targeting tumor angiogenesis is an established strategy for cancer therapy.We identified 131 genes that were differentially upregulated in mTEC.The expression of DEF6 and TMEM176B was upregulated in tumor vessels of human renal cell carcinoma specimens, suggesting that they are potential targets for antiangiogenic intervention for renal cell carcinoma.
Affiliation: Drug Discovery II, DSP Cancer Institute, Dainippon Sumitomo Pharma Co., Ltd, Osaka, Japan.Show MeSH
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Mentions: Excessive angiogenesis occurs through a series of steps including enhanced EC proliferation and migration.36 Therefore, targeting proliferation and/or migration of EC is one of the most attractive and effective strategies for treating angiogenesis-dependent disorders. We reported that mTEC grow faster and migrate better than mNEC.28 These in vitro characteristics of mTEC represent enhanced tumor angiogenesis in vivo and the genes responsible for increased proliferation or migration of mTEC may serve as ideal targets for antiangiogenic therapy. To identify such molecules, we performed loss-of-function screening of the 131 potential TEC markers in Melanoma-EC, one of the mTEC that showed high activity in the proliferation and migration assays.28 We first cotransfected Melanoma-EC with three different sequences of siRNA per gene. Cell proliferation and/or migration were inhibited by >20% compared with mock-transfected cells using siRNA targeted to 44 genes (Fig.2). Subsequently, three different siRNA specific for each of the 44 genes were used to independently transfect Melanoma-EC. We performed migration and proliferation assay using siRNA and finally selected five genes (Def6, Nsg1, Enah, Tmem176b and Pcdhb22; Table3, Fig.3) whose respective siRNA (two or more per gene) inhibited cell proliferation or migration by >30% (Figs2, 4). Cell migration was inhibited by siRNA of Tmem176b, Pcdhb22, Nsg1 and Enah, but cell proliferation was not inhibited. In contrast, cell proliferation was inhibited by Def6 siRNA. These were considered potential regulators of proliferation or migration of mTEC. There was no gene whose two or more siRNA inhibited both cell proliferation and migration by >30% (data not shown). Knockdown of each gene was confirmed using qRT-PCR 48 h after transfection (Fig.4).
Affiliation: Drug Discovery II, DSP Cancer Institute, Dainippon Sumitomo Pharma Co., Ltd, Osaka, Japan.