Abstract 5013: CpG island shore methylation regulates caveolin-1 expression in breast cancer

Abstract

Abstract Caveolin-1 (Cav1), a multifunctional membrane protein, regulates caveolae-dependent lipid trafficking, vesicular transport and signal transduction. Cav1 expression is decreased in breast carcinomas compared to normal tissue, and loss of Cav1 expression promotes mammary tumorigenesis. However, recent reports indicate that Cav1 is overexpressed in a subset of basal-like breast carcinomas and consider it as a basal marker. To investigate the mechanism that results in differential Cav1 expression in breast cancer, we performed genome-wide profiling of DNA methylation using affinity purification and subsequent next-generation sequencing of eluted DNA (MethylCap-seq) on 30 breast cancer cell lines. In breast cancer cell lines displaying low Cav1 expression (“Cav1-low;” n=15, mostly luminal subtype), Cav1 promoter CpG island (CGI) hypermethylation was observed in only four cell lines. In the remaining Cav1-low lines, dense methylation at “CGI shores” (methylation at regions outside of CGI) was seen, while Cav1 promoter CGI methylation was strikingly low or absent. In “Cav1-high” cell lines (n=15 of basal-like subtype), hypomethylation of both promoter CGI and CGI shores was observed. By integrating the methylome data with Cav1 mRNA expression profiles, we found a significant negative correlation between Cav1 CGI shore methylation and Cav1 gene expression. A similar DNA methylation pattern was observed in a panel of antiestrogen-sensitive and -resistant cell lines previously generated by our lab, with hypermethylation at CGI shores for a Cav1-low, tamoxifen-resistant line, and CGI shore hypomethylation in Cav1-high, fulvestrant-resistant lines (basal-like). Treatment of Cav1-low cells with the DNA methyltransferase inhibitor 5-aza-deoxycytidine resulted in demethylation of CGI shores and Cav1 re-expression, further confirming the regulation of Cav1 expression by CGI shore methylation. ChIP-seq results of Cav1-low cells indicated that the Cav1 CGI hypomethylation was due to H3K4me2 occupancy. SiRNA-mediated Cav1 knockdown resulted in a significant inhibition of cell growth and invasion, indicating an important role of Cav1 in supporting breast cancer cell progression. Furthermore, by using an on-line survival analysis tool (Kaplan Meier Plotter), we determined a significant negative correlation between Cav1 levels and overall survival for patients with ERα-negative breast tumors. The results of this study demonstrate that DNA methylation at Cav1 CGI shores is associated with tumor progression, displaying hypomethylation in normal breast epithelial cells, hypermethylation in luminal breast cancer cells, and hypomethylation in basal-like breast cancer cells. In addition, Cav1 CGI shore methylation may represent a novel prognostic factor for ERα-negative, fulvestrant-resistant breast cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5013. doi:1538-7445.AM2012-5013