210 The Development of Distant Metastases in Postmenopausal Woman with Breast Cancer in Association with VEGF Gene Polymorphisms

Background: The epithelial to mesenchymal transition (EMT) is involved in multiple aspects of cancer biology, including tumor metastasis, cancer stem cells, drug resistance, and immunosuppression. TGFb, one of the most studied regulators of EMT, is produced in a latent form that needs to be activated to enable binding to its receptor and subsequent down-stream signaling. Four of the five aVb integrin proteins have been demonstrated to induce TGFb maturation. Cross-talk between TGFb and integrin signaling can also occur downstream of initial receptor activation and regulates various cellular processes, including EMT. Materials and Methods: Lung, melanoma, and breast cancer cell lines were obtained from ATCC. Protein levels were determined by Western-blot and RNA levels were determined by q-PCR. Migration and invasion assays were carried out using trans-well system. All experiments were conducted without coating the plates with vitronectin. Results: Active TGFb1 increased integrin aV, b1, and b3 expression in lung cancer cells while integrin b5 expression was suppressed by TGFb1. This differential regulation of integrin protein expression was accompanied by a reorganization of integrin heterodimer complex between aV and b subunits. TGFb-treated cells developed a spindle-like morphology. Altered expressions of E-Cadherin, vimentin, and snail in these cells were consistent with EMT. Intetumumab (CNTO95), a monoclonal antibody that binds to aVb integrins (aVb1, aVb3, aVb5, aVb6, and aVb8), potently inhibits cellular events strongly relevant to EMT such as migration, invasion and metastasis. Co-treatment of cells with intetumumab blocked TGFb-induced EMT. Moreover, lung cancer cell lines treated with intetumumab alone underwent a mesenchymal to epithelial transition (MET), with increased expression of E-cadherin and decreased expression of vimentin and snail. The effect of intetumumab on EMT was also observed in melanoma cancer cells. Conclusions: aVb integrins are critical regulators of EMT and MET. Intetumumab activates MET and inhibits TGFb induced EMT by blocking aVb integrins, suggesting potential roles in preventing metastasis.