Abstract 2964: Epigenetic mechanisms of transformation by WHSC1L1 in breast cancer cells with the 8p11-p12 amplicon.

Abstract

Abstract An amplified region of chromosome 8, the 8p11-p12 amplicon, is observed in ∼15% of breast cancers. This region contains 55 genes, of which 22 have been identified as candidate oncogenes based on higher than normal copy number and associated elevated expression levels in breast tumor cells compared to normal breast epithelial cells. Our lab previously identified the short isoform of one amplicon gene, Wolf Hirschhorn Syndrome Candidate 1-Like 1 (WHSC1L1), to be a potent inducer of transformation when over expressed in MCF10A breast epithelial cells in vitro. WHSC1L1 is a member of the NSD family of methyltransferases, and has been shown to methylate lysines 4, 27, and 36 of the histone H3 subunit, however, the potently transforming short isoform (WHshort) lacks the catalytic SET domain. Our lab has shown that over expression of WHshort in the immortalized human breast epithelial cell line MCF10A causes a significant increase in proliferation in insulin-free culture conditions, and the ability to form colonies in soft agar. In SUM44 cells, an ER+ breast cancer cell line harboring the 8p11-p12 amplicon, shRNA knockdown of WHSC1L1 resulted in growth arrest. The WHSC1L1 family member NSD1 has been shown to act as an oncogene in acute myelogenous leukemia by blocking differentiation in developing lymphocytes through H3K36me3-mediated inhibition of the polycomb repressor complex, resulting in constitutive activation of the HOXA gene family. While WHSC1L1 may work by a similar mechanism, little is known about the specific substrates of WHSC1L1, or even whether its oncogenic activity is caused by histone methylation changes. Knockdown by shRNA of WHSC1L1 in SUM44 cells resulted in a significant reduction in transcript levels of several genes important in breast cancer cell proliferation, including ESR1, MYCN (amplified in SUM-44), ERBB3, and ERBB4. Interestingly, Carroll et al. recently reported that ER+ breast tumors can be stratified by gene expression signature into poor outcome and good outcome groups, and that ER-alpha binding site patterns on chromatin were altered in the two groups. In this study, WHSC1L1 was part of the ER-regulated gene signature of the poor outcome group. Therefore, we hypothesize that WHSC1L1 may influence histone H3K36me3 marks at specific gene loci, leading to altered ER-alpha binding that is associated with the expression signatures of poor-outcome ER+ breast cancers. To investigate chromatin changes as a result of changes in WHSC1L1 expression, we have accomplished shRNA knockdown of WHSC1L1 in SUM44 cells using GIPZ and TRIPZ lentiviral vectors. Using these cell lines, we are investigating changes in H3K36me3 marks and ER-alpha binding patterns by chromatin immunoprecipitation followed by sequencing (ChIP-seq). These experiments will help to elucidate the interaction of WHSC1L1 and ER in regulated gene expression in breast cancer cells with the 8p11 amplicon. Citation Format: Jonathan C. Irish, Gang Liu, Stephen P. Ethier. Epigenetic mechanisms of transformation by WHSC1L1 in breast cancer cells with the 8p11-p12 amplicon. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2964. doi:10.1158/1538-7445.AM2013-2964