Up to 40% of patients presenting with """"""""superficial"""""""" (i.e. non muscle invasive) bladder cancer develop the muscle """"""""invasive"""""""" life threatening form of the disease during subsequent follow up. In clinical studies, over expression of the epidermal growth factor receptor (EGFR) and the Ras oncogene have been associated with this phenotypic tumor transition. Using a powerful orthotopic invasion assay coupled with gene transfer techniques, the applicant's previous work in human bladder cancer has suggested a causal link between EGFR and Ras gene over expression, and the invasive phenotype. However the molecular pathways by which these two genes effectively trigger or facilitate the invasive process is incompletely understood. Preliminary and published data implicate Ras and phosphotidylinositol 3-kinase (PI3K) in this process and suggest that these molecules may carry EGFR mediated signals downstream to Rho family GTP binding proteins, which in turn control cellular motility and hence contribute to the invasive phenotype. The applicants propose to continue these studies and begin to elucidate the EGF mediated pathways leading to enhanced tumor motility in vitro and the impact this has on tumor invasion in vivo, by testing the following hypotheses: 1) EGFR over expression can confer the invasive phenotype to superficial bladder cancer, 2) when EGFR confers the invasive phenotype, this is mediated via both Ras and PI3K, 3) activation of Rho family proteins is necessary in order for EGFR mediated signals to induce invasion and serve to integrate signals from both Ras and PI3K in this process. To test these hypotheses they propose the following Specific Aims: 1) Alter the levels of EGFR in bladder cancer cell lines by over expressing wild type EGFR in superficial cell lines and suppressing EGFR expression in invasive cell lines, 2) Modulate PI3K and Ras activity in cells over expressing EGFR by using dominant negative and constitutively active PI3K and Ras constructs and 3) Determine if Rho family GTP binding proteins are effectors of EGF induced motility and invasion. They conclude that because their orthotopic assay closely mimics the behavior of bladder cancer in humans, successful completion of these specific aims will provide biologically relevant molecular information on the signalling pathways regulating EGF stimulated bladder cancer invasion in vivo and may provide a foundation for the development of novel therapies that interfere with this process in patients with superficial bladder cancer.
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