Abstract P5-06-01: Distinctive coding and non-coding RNA profiles of pre-menopausal and post-menopausal benign breast
Authors: Carter JM , Nair AA , Davila JI , Heinzen EP , Hoskin TL , Winham SJ , Radisky DC , Degnim AC , Visscher DW 
Published: May 2019

Background: Despite a paucity of data on the benign breast transcriptome, reduction mammoplasty tissue is commonly used as a control for RNA biomarker discovery in breast cancer (BC). We evaluated the transcriptome of benign breast to characterize pre-menopausal and post-menopausal coding and non-coding RNA profiles. These profiles will provide insight into the metabolic landscape of benign breast and inform transcriptomic studies of BC and peri-tumoral microenvironments.

Methods: With institutional approval, cryobanked fresh breast tissues from reduction mammoplasties (age 15-62y; follow-up: 4-10 y with no cancer events), with histologically-confirmed benign tissues with similar epithelial: stromal tissues were grouped into 4 age sets (&lt; 30y, N=10; 30-39y, N=11; 40-49y, N=10; ≥50y (clinically post-menopausal), N=10). Following RNA extraction, library preparation (Illumina TruSeq Stranded mRNA kit), and sequencing (Illumina HiSeq 4000), reads were processed (MAP-RSeq v3.0.0) and aligned (STAR aligner; hg38). Differential expression (DE) analysis (edgeR 2.6.2) identified DE genes from normalized RPKM reads (absolute log2 fold change (FC) &gt; 1 and false discovery rate (FDR) &lt; 0.10), corrected for intra-group biases using medians (absolute FC cut-off of &gt;1.5). Over-representation analysis [Ingenuity pathway analysis (IPA), Ingenuity® Systems] and gene set enrichment analysis [(GSEA), GeneTrail 2.0] identified significantly-enriched pathways.

Results: Across the 4 age sets, 561 DE genes were identified. Compared to the post-menopausal (PM) set, the number of DE genes was highest in &lt;30 y set (N= 372) and decreased with increasing age (N= 170, age 30-39 set and N=20, age 40-49 set), generating up-regulated (PMup) and down-regulated (PMdown) PM transcriptomic profiles. The top PMdown DE genes included RANKL, WNT4, MKI67, extracellular matrix, and lactation-related genes (lactoferrin ,MUC4, MUC16, p &lt; 0.01). Top PMdown canonical pathways were cell cycle-related (CDK1, CCNA2, CCNB2, ESPL1, TOP2A)(p&lt; 0.001). Top PMup genes included those involved in adipogenesis, NPY1R, NPY2R, unsaturated fatty acid synthesis and eicosanoid signaling (P&lt; 0.001). Top PMup canonical pathways included acyl co-A hydrolysis, stearate biosynthesis, acute phase response and RXR signaling (P &lt; 0.001). GSEA of the entire gene set (N=15,466; ranked in order of 561 DE genes) identified 16 significant pathways, functionally grouped as mitochondrial energy metabolism, proteasome, and unsaturated fatty acid biosynthesis and signaling (PPAR pathway). While coding RNAs comprised 85% of DE transcripts, lncRNA comprised ˜5%. Top DE lncRNAs and precursor miRNAs included MEG3, a tumor suppressor, LIN00092, miR22 and miR1182 (p &lt; 0.0001).

Conclusions: Pre and post-menopausal benign breast tissues have distinctive transcriptomic profiles. PM benign breast has down-regulated proliferation/cell-cycle-related pathways, and up-regulated genes in mitochondrial energy pathways, lipogenesis and inflammatory pathways. Notably, many of the top DE coding and non-coding RNAs in PM benign breast have been implicated in BC progression, highlighting their value in better understanding breast carcinogenesis and the need to characterize their functional roles in normal aging and menopause.

Citation Format: Carter JM, Nair AA, Davila JI, Heinzen EP, Hoskin TL, Winham SJ, Radisky DC, Degnim AC, Visscher DW. Distinctive coding and non-coding RNA profiles of pre-menopausal and post-menopausal benign breast [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-06-01.