Rosen, Kirill
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Item Open Access Oncogenic ras-induced downregulation of a pro-apoptotic protease caspase-2 is required for malignant transformation of intestinal epithelial cells(Journal of Biological Chemistry, 2011-11-11) Yoo, Byong Hoon; Rosen, Kirill V.;Resistance of carcinoma cells to anoikis, apoptosis that is normally induced by loss of cell-to-extracellular matrix (ECM) adhesion, is thought to be essential for the ability of these cells to form primary tumors, invade adjacent tissues and metastasize to distant organs. The current knowledge about the mechanisms by which cancer cells evade anoikis is far from complete. In an effort to understand these mechanisms we found that ras, a major oncogene, downregulates protease caspase-2 (that initiates certain steps of the cellular apoptotic program) in malignant human and rat intestinal epithelial cells. This downregulation could be reversed by inhibition of a protein kinase Mek, a mediator of Ras signaling. We also found that enforced downregulation of caspase-2 in non-malignant intestinal epithelial cells by RNA interference protected them from anoikis. Furthermore, the reversal of the effect of Ras on caspase-2 achieved by the expression of exogenous caspase-2 in detached ras-transformed intestinal epithelial cells promoted well established apoptotic events, such as the release of the pro-apoptotic mitochondrial factors cytochome c and HtrA2/Omi into the cytoplasm of these cells, significantly enhanced their anoikis susceptibility and blocked their long-term growth in the absence of adhesion to the ECM. Finally, the blockade of the effect of Ras on caspase-2 substantially suppressed growth of tumors formed by the ras-transformed cells in mice. We conclude that ras-induced downregulation of caspase-2 represents a novel mechanism by which oncogenic Ras protects malignant intestinal epithelial cells from anoikis, promotes their anchorage-independent growth and allows them to form tumors in vivo.Item Open Access Tumor suppressor protein kinase Chk2 is a mediator of anoikis of intestinal epithelial cells(International Journal of Cancer, 2011-11-02) Yoo, Byong Hoon; Rosen, Kirill V.Resistance of carcinoma cells to anoikis, apoptosis that is normally induced by detachment of non-malignant epithelial cells from the extracellular matrix, is thought to be critical for carcinoma progression. Molecular mechanisms that control anoikis of non-malignant and cancer cells are understood poorly. In an effort to understand them we found that detachment of non-malignant intestinal epithelial cells triggers upregulation of Chk2, a pro-apoptotic protein kinase that has never been implicated in anoikis and has been thought to kill cells mainly under the conditions compromising genome integrity. We found that enforced downregulation of Chk2 protects intestinal epithelial cells from anoikis. Chk2 can kill cells by stabilizing p53 tumor suppressor protein or via p53-independent mechanisms, and we established that Chk2-mediated anoikis of intestinal epithelial cells is p53-independent. We further found that, unlike non-malignant intestinal epithelial cells whose anoikis is triggered by detachment-induced Chk2 upregulation, intestinal epithelial cells carrying oncogenic ras, a known inhibitor of anoikis, remain anoikis-resistant in response to enforced Chk2 upregulation. By contrast, drugs, such as topoisomerase I inhibitors, that can kill cells via Chk2-indpendent mechanisms, efficiently triggered anoikis of ras-transformed cells. Thus, oncogenic ras can prevent Chk2 from triggering anoikis even when levels of this protein kinase are elevated in cancer cells, and the use of therapeutic agents that kill cells in a Chk-2-independent, rather than Chk-2-dependent, manner could represent an efficient strategy for overcoming ras-induced anoikis resistance of these cells. We conclude that Chk-2 is an important novel component of anoikis-promoting machinery of intestinal epithelial cells.