Abstract A25: Genome-wide methylation analyses in advanced-stage prostate cancer models

Abstract

Abstract Introduction: Aberrant DNA methylation is a frequent event in prostate cancer (PCa). Investigation of genome-wide methylation changes in PCa promises to provide novel biomarkers that discriminate between indolent and aggressive diseases and to facilitate the development of novel treatment options. Most methylation studies to date have focused on CpG islands (CGI) in the promoter region of known genes. The current study investigated genome-wide methylation changes with the aim to identify recurrent regions differentially methylated in advanced PCa, including metastases and castration-resistance diseases. Methods: Using comprehensive high-throughput array-based relative methylation analysis (CHARM), we assessed genome-wide methylation status in metastatic PCa represented by two PCa cell lines (LNCap and VCap) as well as castration-resistant cancer represented by PCa xenografts cultivated in castrated mice (LuCaP23.1v, 35v, and 96v) and APIPC cells derived from AR-null LNCap maintained in hormone depleted media. To identify differentially methylated regions (DMR), each metastatic PCa cell line was compared with benign prostate epithelial RWPE1 cells. The castration-resistant xenografts were compared with the corresponding castration-sensitive xenografts, LuCaP23.1, 35, and 96, respectively. APIPC was compared with the parent line maintained in regular growth media. Unlike most methylation analyses that only target promoter CGI, CHARM analyses target CpG enriched regions of the entire genome, including genomic sequences up to 2kb outside of promoter CGIs, “CpG island shores”. The shores have been recently demonstrated to harbor more methylation alterations than the islands in tissue differentiation and cancer development. Results: Methylation changes were frequent in PCa metastases and castration-resistant diseases. Using RWPE1 as the reference, 306 DMRs were identified in LNCap (114 hyper- and 192 hypo-methylated), while 128 DMRs (32 hyper- and 96 hypo-methylated) were discovered in VCap cells. Compared with the parent LNCap line, APIPC showed 284 DMRs (80 hyper- and 204 hypomethylated). Using each corresponding castration-sensitive xenograft as the reference, castration-resistant LuCaP23.1v, LuCaP35v, and LuCaP96v demonstrated 587 (378 hyper- and 209 hypo-methylated), 150 (17 hyper- and 133 hypo-methylated), and 761 (353 hyper- and 408 hypo-methylated) DMRs, respectively. Distribution analyses showed that DMRs associated with these advanced PCa are enriched in regions 500bp outside CpG islands, i.e., CpG island shores, rather than in the islands. As expected, recurrent DMRs were evident. LNCap and VCap, representing PCa metastases, demonstrated 20 DMRs in common. Two of these common DMRs were of special interest: one is located at the 3′ UTR of STAT6, the expression of which was previously shown to correlate with high histological grades of prostate cancer and with tumor size; another DMR is located at intron 1 of HOXA9 within the CGI shore. Aberrant methylation of HOXA9 promoter CGI was reported in multiple cancers such as bladder and non-small cell lung cancer. Among those representing castration-resistant PCa, 11 DMRs were shared between APIPC and LuCaP23.1v, while 2 DMRs were in common between APIPC and LuCaP96v. Conclusion: CHARM studies in advanced prostate cancer enabled us to better understand methylation changes at the whole genome level. Our data confirmed the importance of CpG enriched regions outside the promoter CGI during the development of cancer, specifically, PCa metastases and castration-resistant diseases. Multiple novel recurrent DMRs were identified in the study, which call for further investigation of their role in primary PCa as well as the related changes in gene expression and histone modification, and the prognostic significance of those changes. Citation Format: Xiaoyu Qu, Jerry Davison, Eric Bluemn, Peter Nelson, Robert Vessella, Min Fang. Genome-wide methylation analyses in advanced-stage prostate cancer models [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr A25.