Sleeping Beauty mediated targeting of tumor endothelial marker 8 in the tumor vasculature of colorectal carcinomas in mice

Recent advances in cancer research have broaden the knowledge of how solid tumors progress. Once thought of as the product of a single aberrant cell with multiple mutations, tumors are the product of that single cell dividing, and then the new mass interacting with multiple nonmalignant cell types that guide its growth. Some studies examined the expression profiles of the nonmalignant cell types that associate with tumors and compared their profile to the same cell type in normal tissues. The results showed that the cells associating with the tumors expressed abnormal proteins.;Tumor Endothelial Marker 8 (TEM8) is a recently described protein preferentially expressed on tumor associated endothelial cells. The role played by TEM8 in endothelial biology remains unclear, yet this protein may allow specific delivery of therapy to tumor endothelium. This research describes the creation of a fusion protein that specifically targets TEM8 to deliver a potent vascular disrupting agent to tumor vessels.;Western blot analysis, enzyme-linked immunosorbent assays and enzymatic assays evaluated the specificity and function of the fusion proteins in vitro. In vivo analysis utilized a xenograft model of colorectal carcinoma to test the efficacy of targeted and control fusion proteins after gene delivery with Sleeping Beauty. Tumor growth curves as well as survival studies evaluated animals receiving gene delivery of the various fusion proteins. Histology and immunohistochemistry visualized the expression and homing of the fusion proteins to the tumor vasculature, as well as effects on tumor vessel density and thrombosis.;In vitro analysis confirmed the predicted antigen specificity and function of the fusion proteins. Mice treated with the gene encoding anti-TEM8/tTF exhibited a 55% reduction in tumor volume when compared to the untreated animals (p < 0.001) and achieved a 49% increase in tumor growth delay by Kaplan Meier analysis (p = 0.0367). Immunohistochemistry confirmed tumor endothelial expression of TEM8, fusion protein homing to tumor vasculature, a decrease in vessel density and localized areas of thrombosis. Collectively, the results presented in this study indicate that targeting TEM8 is a viable anti-tumor strategy that deserves further consideration and development.