Abstract LB-378: Transcriptome analysis of tamoxifen-resistant breast cancer cells using next generation sequencing

Abstract

Abstract Resistance to tamoxifen (Tam), a widely used antagonist of the estrogen receptor (ER), is a common problem in breast cancer therapy. At least 70% of breast cancers are ER positive, constituting eligible candidates for Tam treatment. Nonetheless, recurrent disease occurs in one third of patients treated with Tam within 15 years of therapy, implying that approximately one-quarter of all breast cancers are Tam-resistant either in initial presentation or recurrent disease. Thus, Tam resistance has become a major clinical issue in the treatment of breast cancer. Yet, the molecular mechanism behind Tam resistance is still not completely understood. A better understanding of the genetic or epigenetic changes associated with Tam resistance should facilitate circumventing this problem. In this study, using next generation sequencing technology, we sought to compare the transcriptomes of Tam sensitive and resistant breast cancer cells, in order to identify genes involved in the development of Tam resistance. rRNA depleted total RNA from MTR-3 (a Tam-resistant breast cancer cell line derived by culturing MCF-7 cells continuously for a two-year period in the presence of 1 μM Tam) and from parental MCF-7 cells were sequenced on an Applied Biosystems-SOLiD V3 sequencer. Short-read (50 bp) libraries were sequenced and mapped to the reference human genome (hg18) by Maq, and whole transcriptomes for the two cell lines were constructed and compared for their gene expression. 140 million total reads were produced by sequencing, and ∼ 50% were accurately mapped. AB WT Analysis Pipeline was used to confirm results and score expression. Gene network and pathway analysis were constructed using Ingenuity Pathway Analysis (IPA) and GeneGeo. Potential gene candidates were validated using TaqMan Gene Expression assays. Based on a 1.7 fold criterion, we found that 667 genes, including those vital to cell cycle, cell death, cancer development, and energy metabolism, were significantly up- or down- regulated. Gene network analysis indicates that the most enriched pathways are related to ERα. Notably, Sirtuin 3 (SIRT3), a gene encoding a mitochondrial deacetylase important in cellular energetic and stress responses, was implicated in the next generation sequencing data and confirmed by qPCR to be up-regulated in the Tam-resistant cells, suggesting that SIRT3 is a potential important player in the regulation of tamoxifen resistance. The results of the current study may uncover previously unrecognized genes and molecular pathways that contribute to Tam resistance. Analysis of the deep sequencing data, identification, and validation of other potential Tam resistance-associated genes are still in progress. Our study suggests that comparative analysis of transcriptomes has the potential to help identify new targets that can be exploited to modulate sensitivity of breast cancer cells to this clinically useful anti-estrogen agent. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-378. doi:10.1158/1538-7445.AM2011-LB-378