) Anchorage-dependence is one of the best in vitro correlates of tumor cell malignancy in vivo. Several years ago, our laboratory discovered that normal epithelial cells undergo apoptosis if appropriate integrin-mediated matrix contacts are lost (""""""""anoikis""""""""), which re-defines epithelial anchorage dependence.The present study will focus on elucidating a novel mechanism by which death receptors (e.g., TNFR1 and FAS) regulate anoikis, anchorage-dependence, and, by extension, tumor cell malignancy. Alterations in death receptor-related proteins are observed in several tumor cell systems, suggesting their importance in human cancer. However, their specific role in this regard is unclear at present. Recently, we have found evidence that death receptors may be involved in initiating anoikis. Overall, the role of death receptors and their mechanism of activation in anoikis, and how these mechanisms are regulated by oncogenes, will be examined in this project. We will address the following questions in particular. First, are the death receptors themselves activated (i.e., in terms of TNFR1 clustering and TNFR1-TRADD association) in cells detached from extracellular matrix? If so, do signaling molecules that are known to regulate anoikis (CD2-FAK, activated ras, Akt or c-src) affect receptor activation? Secondly, does death receptor activation in anoikis occur independently of a death ligand? (e.g., by a cytoskeletal sequestration vs. release mechanism, or by modification of death receptor signaling components) or does it occur due to the de novo production of a death ligand? (e.g., translational upregulatation or proteolytic processing). Thirdly, what is the role of cadherin-catenin complexes in the regulation of anoikis? and can these complexes regulate death receptor activation? In summary, this project will establish the basic parameters underlying the activation of death receptors in cells deprived of normal matrix interactions. It will also reveal novel mechanisms wherein oncoproteins or cell adhesion molecules can regulate anoikis. It is anticipated that these studies will reveal novel mechanisms by which tumor cells become defective in anoikis and how these defects might be compensated by gene therapy or drugs.
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