We found that BT474 cells express detectable levels of Puma and of Bim whether cells were grown under con trol conditions or transfected with control, scramble siR NAs. In contrast, these cells selleck kinase inhibitor expressed barely detectable levels of Noxa, a BH3 only protein which functions as a selectiove inhibitor of Mcl 1. Regarding Bim, it has to be noted that we essentially detected its Bim Extra Long form, whereas the Long and Short forms were less expressed in these cells. To investigate whether Bim or Puma play an active role in the Mcl 1 dependence of BT474 cells, these cells were transfected with control, Bim or Puma siRNA, which down regulated efficiently the targeted proteins, prior to their transfection with Mcl 1 siRNA and investigation of cell death. Of note, neither Bim nor Puma siRNA affected cell viability by themselves.

Bim depletion robustly prevented cell death induced by transfection with Mcl 1 siRNA, as measured by APO2. 7 staining or by Annexin V staining, indicating that this pro apoptotic protein plays a major role in the Mcl 1 dependence of BT474 cells. In contrast, PUMA depletion had a much Inhibitors,Modulators,Libraries less pronounced and consistent effect on Mcl 1 knock down induced cell death. We investigated whether Bim contributes to the Mcl 1 dependence of the subpopulation of BT474 that are cap able of forming mammospheres. Bim depletion had no impact in itself on mammosphere formation by BT474 cells. However, it abrogated the ability of Mcl 1 knock down to decrease the number of mammospheres formed by BT474 cells. This is strong support to the notion that the Mcl 1 dependence of BT474 CICs also is due to Bim expression.

It rises from above that Inhibitors,Modulators,Libraries constitutive expression of Bim may contribute Inhibitors,Modulators,Libraries to render Mcl 1 necessary for the survival of HER2 overexpressing tumors. To analyze whether mechanisms leading to Bim transcription are particularly at stake in HER2 overexpressing tumors, we went back to our investigation of published gene expres sion profiles of breast cancer patients using a probe matching approach as described above. As shown in Table 1, we found a statistically significant enrichment, in HER2 overexpressing breast tumors compared to other breast tumors, in one BCL2L11 specific probe. Regarding pro apoptotic genes, a statistical enrichment in one BID specific probe and in one BIK specific probe was also found.

In contrast, other breast tumors appeared statistically enriched for two PMAIP1 specific probes and for one BAD specific one. While this tends to suggest that pathways leading to Bim transcription might Inhibitors,Modulators,Libraries be more active in HER2 overexpressing breast cancers, this should nevertheless be taken cautiously. Indeed, we did not confirm a statistical enrichment for Bim expression in Inhibitors,Modulators,Libraries HER2 overexpressing cancers by our gene http://www.selleckchem.com/products/Bortezomib.html matching approach involving 5 cohorts, even though enrichment for BID and BIK and impoverish ment for BAD and NOXA were confirmed.