Screening for Small Molecules That Disrupt IAP-Caspases Binding to Activate Caspases and Induce Apoptosis in Breast Cancers
Final rept. 1 Sep 2004-1 Aug 2005
MICHIGAN UNIV ANN ARBOR
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IAP Inhibitor of Apoptosis Protein suppresses apoptosis through binding and inhibiting active caspases-3, -7 and -9 via its BIR baculoviral lAP repeat domains. During apoptosis the caspase inhibition by lAPs can be negatively regulated by a mitochondrial protein Smac. Smac physically interacts with multiple lAPs and relieves their inhibitory effect on caspases-3, -7 and -9. Recently, a small molecule Smac-mimic compound Smac-mimic, which potentiates TRAIL and TNF-a mediated cell death in glioblastoma T98G cells and HeLa cells, was identified and characterized. To determine the efficacy of this compound in breast cancer cells, we first measured protein expression of three IAPs XIAP, cIAP-i, and cIAP -2 in 9 independent breast cancer cell lines. Three cell lines were chosen a high IAPs expressing line MDA-MB- 231, and two low lAPs expressing lines, T47D and MDA-MB-453. The cell lines were tested for their sensitivity to Smac-mimic alone or in combination with TRAIL or etoposide. Acting alone, Smac-mimic was quite potent with a cytotoxic 1C50 of 3.8 nM in high IAPs expressing MDA-MB-231 cells, but was inactive at a much higher concentration in low IAPs expressing T47D and MDA-MB-453 cells. In fact, as low as 2.5 nM of Smac-mimic alone was sufficient to activate caspase-3 and induce apoptosis in MDA-MB-231 cells. In combinational treatments with TRAIL or etoposide, Smac-mimic significantly sensitized cells to growth suppression in MDA-MB-231 cells, but to a lesser extent, in T47D and MDA-MB-453 cells. Furthermore, it significantly synergized MDA-MB-231, but not T47D cells to apoptosis induced by either TRAIL or etoposide. Thus, in these cell lines, Smac-mimic acts in an apparent lAPs dependent manner to induce apoptosis alone as well as sensitizes breast cancer cells to TRAIL or etoposide induced apoptosis via caspase 3 activation.
- Medicine and Medical Research