Abstract #3272: Regulation of Mcl-1 and its contribution to cell survival and drug resistance in breast cancer

Mcl-1 is a tightly regulated member of the Bcl-2 family that plays an important role in regulating the balance between life and death in a variety of cell types. Levels of Mcl-1 fluctuate dramatically in response to activated signalling pathways within the cell. Over-activation of survival signalling can confer a survival advantage by upregulating survival proteins such as Mcl-1. In breast cancer, signalling via the epidermal growth factor receptors is frequently deregulated and increased levels of Mcl-1 have been correlated with elevated expression of key EGF receptors. Another pathway frequently exploited by breast tumour cells is signalling via estrogen receptor alpha. We aim to investigate the relationship between these pathways and Mcl-1 gene expression. We hypothesize that both pathways transcriptionally elevate Mcl-1 levels, which leads to increased survival and resistance to apoptosis. Knowledge gained from this study will determine whether specific inhibition of Mcl-1 activity may have a potential therapeutic benefit in this type of cancer. We have found that treatment of breast cancer cell lines with estrogen and Epidermal Growth Factor (EGF) led to increased resistance to chemotherapeutic drugs and coincided with elevated Mcl-1 protein levels in breast cancer cell lines. Knock-down of Mcl-1 by shRNA or inhibition of its action through small molecule inhibitors reduced viability and sensitized cells to drug induced apoptosis. A dramatic increase in mRNA levels was detected by real time PCR following EGF stimulation suggesting that the increase in protein is likely a result of enhanced transcription. Elevated levels of Mcl-1 mRNA were also observed upon treatment with estrogen. In silico promoter analysis indicates the presence of a variety of transcription factor binding sites, including half estrogen responsive elements that may be involved in regulation of expression. Transcriptional regulation of Mcl-1 was further studied through cloning of the full-length Mcl-1 promoter and preparation of specific deletion mutants followed by luciferase assays. The critical region for EGF induction was narrowed down to a small fragment containing a predicted Nf-\#312;B binding site. Chromatin immunoprecipitation as well as electromobility shift assays will be employed to assess specific transcription factor binding to the Mcl-1 promoter. These results indicate that Mcl-1 plays an important role in promoting cell survival and resistance to apoptosis in breast cancer. Combination of Mcl-1 inhibitors with traditional chemotherapy may overcome resistance conferred by over-activation of survival signalling pathways.

Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 3272.