Abstract 186: Methyl binding domain protien 2 (MBD2) promotes epithelial dedifferentiation in breast cancer

Abstract

Abstract Purpose: Methyl-CpG Binding Proteins (MCBPs) are thought to function as the interpreters of epigenetic information encoded in cytosine methylation. Their ability to translate DNA methylation into local transcriptional repression has sparked interest in the role MCBPs play in cancer. We have studied the function of Methyl-Binding Domain Proteins (MBDs) in human mammary epithelial cancers, where repatterning of CpG methylation is common. Results: We find Methyl Binding Domain Protein 2 (MBD2) promotes the abnormal multi-cellular morphology characteristic of tumor cells. Stable MBD2 knockdown in MCF7 cells leads to an increased proportion of differentiated epithelial structures (e.g. acinii, 70%, [CI=0.55-0.83]) in 3D culture when compared to untransfected (46%, [CI=0.39-0.53], p ≤ 0.038) or control shRNA transfected (37%, [CI=0.29-0.45], p ≤ 0.012) populations. To identify the genes underlying this MBD2 dependant phenotype, high throughput quantitative PCR data were probed using self organizing map (SOM) analysis. We found only a small subset of the breast cancer specific tumor suppressors known to be silenced by promoter hypermethylation were regulated by MBD2 alone (n=7, 15%). Several genes previously shown (by us and others) to be MBD2 bound by chromatin immunoprecipitation (ChIP) were not induced by MBD2 knockdown, suggesting MBD2 exhibits tumor and tissue-type specificity. Moreover, gene sets activated by the DNA methylation inhibitor, 2-deoxy-5-aza-cytidine (4uM, n=30, 67%), and MBD2 knockdown (n=18, 40%) were different, suggesting pleiotropic activity; for example at loci that exhibit additive induction (n=11, 24%) and those that do not. MBD2 dependant genes were rapidly re-suppressed upon rescue with a shRNA binding site variant MBD2 and ChIP studies confirmed binding of MBD2 at all genes examined within the MBD2 dependant cluster. Transient siRNA mediated knockdown of individual MBD2 dependant genes did not restore the dedifferentiated phenotype, suggesting coordinate regulation by multiple downstream targets of MBD2. Conclusion: Our studies show MBD2 promotes tumor suppressor silencing and dedifferentiation in breast cancer and intimate a role for MBD2 in fine-tuning mammary epithelial gene expression patterns. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 186.