Abstract P2-04-02: Comparison of DNA methylation patterns in normal breast tissue from women with and without breast cancer
Authors: Hofstatter EW , Zhu Y , Horvath S , Chagpar AB , Wali VB , Bossuyt V , Storniolo AM , Hatzis C , Patwardhan G , Von Wahlde M-K , Butler M , Epstein L , Stavris K , Sturrock T , Au A , Kwei S , Pusztai L 
Published: February 2018

Abstract
BACKGROUND: Increasing evidence suggests that epigenetic mechanisms play critical roles in the development of breast cancer. However, precise DNA methylation signatures associated with breast cancer susceptibility remain unknown. We sought to compare DNA methylation changes in the normal breast tissue of women with and without breast cancer to identify patterns of aberrant DNA methylation in women with breast cancer.
METHODS:Samples of normal breast tissue were collected from four cohorts of women: age &lt; 50 years with and without breast cancer, and age ≥50 years with and without breast cancer. Normal breast tissue from healthy women was obtained from the Komen Tissue Bank at IU Simon Cancer Center and from women presenting for reduction mammoplasty at Yale New Haven Hospital. Normal breast tissue from women with breast cancer was obtained from patients undergoing adjuvant total mastectomy at Yale Breast Center. DNA was extracted using Qiagen AllPrep Universal kit. Raw data files in idat format were imported to Partek Genomics Suite 6.6 for normalization and differential methylation analysis. Raw intensities were normalized using With Array Normalization (SWAN) method. Principal component analysis (PCA) were performed as quality control. Differentially methylated loci (DML) between control and breast cancer groups were detected when False discovery rate (FDR) &lt; 0.05 and fold change &gt; 1.5. Functional enrichment analysis of genes with DML in the gene body were conducted using METACORE™. Pathways with FDR &lt; 0.05 were selected.
RESULTS: Ninety-three normal breast tissue samples from 89 subjects were analyzed (breast cancer=40, unaffected=53). Comparison of DNA methylation patterns between women with and without breast cancer revealed 200 DMLs. The majority of DMLs (186) were hyper-methylated in breast cancer patients, and 48 DMLs locate in enhancers of genes. 170 DMLs locate in 134 genes, enriched in two pathways: (1) Cell adhesion_Endothelial cell contacts by junctional mechanisms, and (2) Neurophysiological process_Constitutive and regulated NMDA receptor trafficking. Genes associated with cell adhesion and cell contacts included: ACTN2, GJA4, GJA7 and MAGI1. Two hyper-methylated loci were found in enhancers of ACTN2. In addition, one hyper-methylated locus in GJA4, one hyper-methylated and one hypo-methylated loci in GJA7, and two hyper-methylated loci in MAGI1 were detected in breast cancer patients. Genes associated with NMDA receptor trafficking include: TPK1, ADCY4 and LIN7C. One and two loci were found in TPK1 and ADCY4, respectively, that were hyper-methylated in normal breast tissue from cancer patients in the gene body, while a hypo-methylated locus in breast cancer patients was identified in LIN7C.
CONCLUSIONS: Comparison of DNA methylation patterns of normal breast tissue from women with and without breast cancer reveal specific mechanistic pathways and genes that are differentially methylated in women with breast cancer. DNA methylation of normal breast tissue deserves further study as a potential biomarker for breast cancer risk stratification and may lend new insight into mechanisms of breast cancer development.
Citation Format: Hofstatter EW, Zhu Y, Horvath S, Chagpar AB, Wali VB, Bossuyt V, Storniolo AM, Hatzis C, Patwardhan G, Von Wahlde M-K, Butler M, Epstein L, Stavris K, Sturrock T, Au A, Kwei S, Pusztai L. Comparison of DNA methylation patterns in normal breast tissue from women with and without breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P2-04-02.