Citations

1501 Background: Despite BC risk reduction of 50-65% by preventive endocrine therapy (ET), very few at-risk women choose to take them. A woman’s perceived BC risk correlates with uptake of ET. A PRS comprised of 77 BC genetic susceptibility loci (Single Nucleotide Polymorphisms (SNP) improves the accuracy of risk prediction for BC. We examined the impact of the addition of individualized PRS BC risk prediction to standard risk calculator estimates on intent to take BC prevention medication. Methods: Eligible women had ≥5% 10 yr BC Tyrer-Cuzick risk (IBIS) or 5 year Gail score ≥3%, with no history of BC or hereditary BC syndrome. Standard BC risk estimates (IBIS or Gail) were incorporated into the counselling on BC preventive ET. A self-reported questionnaire at baseline quantified intention to take ET and explored factors associated with this decision. Blood samples were obtained and genotyped for 77 SNPs, individualized PRS were calculated then incorporated into IBIS and Gail predictions for 5 yr, 10 yr, & lifetime BC risk. At a second visit, PRS risk & prevention recommendations were revisited. Post visit questionnaires assessed change in intent to take ET. Multivariable linear regression was performed to assess impact of baseline variables on change in intent to take medication. Results: From 2016 to 2017, 151 women in Canada & USA were enrolled, median age: 56.1 (range 36-76.4), 35.6% were premenopausal, 98.7% were Caucasian. Median 5yr, 10yr, & lifetime IBIS risk estimates were 3.8% (2.0-11.5), 7.9% (5.0-23.1), and 25.3% (5.5 to 92.2). PRS increased BC risk estimates in 84 (55.6%) and reduced BC risk estimates in 67 (44%) women. After PRS risk counselling, intention to take ET significantly changed (p<0.001): 41.9% of those with increased PRS were more inclined, and 46.7% of women with decreased PRS were less inclined to take ET. On multivariable regression, increase in PRS (p<0.0001) and less concern about ET side effects (p<0.0001) were associated with greater intent to take ET. Conclusions: In high risk women, PRS significantly changed BC risk estimates & intent to take preventive ET. Further assessments of the impact of PRS scores on compliance with ET are warranted. Clinical trial information: NCT02517593.

Abstract Background Archived formalin fixed paraffin embedded (FFPE) samples are valuable clinical resources to examine clinically relevant morphology features and also to study genetic changes. However, DNA quality and quantity of FFPE samples are often sub-optimal, and resulting NGS-based genetics variant detections are prone to false positives. Evaluations of wet-lab and bioinformatics approaches are needed to optimize variant detection from FFPE samples. Results As a pilot study, we designed within-subject triplicate samples of DNA derived from paired FFPE and fresh frozen breast tissues to highlight FFPE-specific artifacts. For FFPE samples, we tested two FFPE DNA extraction methods to determine impact of wet-lab procedures on variant calling: QIAGEN QIAamp DNA Mini Kit ("QA"), and QIAGEN GeneRead DNA FFPE Kit ("QGR"). We also used negative-control (NA12891) and positive control samples (Horizon Discovery Reference Standard FFPE). All DNA sample libraries were prepared for NGS according to the QIAseq Human Breast Cancer Targeted DNA Panel protocol and sequenced on the HiSeq 4000. Variant calling and filtering were performed using QIAGEN Gene Globe Data Portal. Detailed variant concordance comparisons and mutational signature analysis were performed to investigate effects of FFPE samples compared to paired fresh frozen samples, along with different library preparations. In this study, we found that five times or more variants were called with FFPE samples, compared to their paired fresh-frozen tissue samples even after applying molecular barcoding error-correction and default bioinformatics filtering recommended by the vendor. We also found that QGR as an optimized FFPE-DNA extraction approach leads to much fewer discordant variants between paired fresh frozen and FFPE samples. Approximately 92% of the uniquely called FFPE variants were of low allelic frequency range (<5%), and collectively shared a “C>T|G>A” mutational signature known to be representative of FFPE artifacts resulting fromcytosine deamination. Based on control samples and FFPE-frozen replicates, we derived an effective filtering strategy with associated empirical false-discovery estimates. Conclusions Through this study, we demonstrated feasibility of calling and filtering genetic variants from FFPE tissue samples using a combined strategy with molecular barcodes, optimized DNA extraction, and bioinformatics methods incorporating genomics context such as mutational signature and variant allelic frequency.

Abstract 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.

Abstract Background: Differences in ER+ and ER- breast cancer tumor biology are well-documented, but little is known about the field of background benign breast in which those cancers arise. We evaluated the transcriptome of benign breast tissues from women with concurrent ipsilateral ER+ and ER- breast cancers (BC) to characterize coding and non-coding RNA profiles. This pilot study provides insight into the transcriptomic landscapes of benign breast tissues in patients with BC, and the microenvironment of the at-risk benign tissue. Methods: With institutional approval, cryobanked breast tissues from patients with concurrent ipsilateral ER+ BC (benign ER+BC, N=14) or ER- BC (benign ER-BC, N=10), were selected for benign tissues with similar epithelial:stromal ratios and grouped into pre (preM) and post-menopausal (PM) groups. Following RNA sequencing (Illumina TruSeq Stranded mRNA kit & 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. Over-representation analysis [Ingenuity pathway analysis (IPA), Ingenuity® Systems] and gene set enrichment analysis [(GSEA), GeneTrail 2.0] identified significantly-enriched pathways. Results: In the PM group, there were 144 DE transcripts between benign ER+BC and benign ER-BC, including coding RNAs (40%), antisense RNAs (35%) and lncRNAs (7%). In contrast, the preM group had no significantly DE genes between benign ER+BC and benign ER-BC. In the PM DE coding gene set, the top DE transcripts in benign ER+BC included many genes implicated in BC development# or ER+ BC progression (*) (e.g. up-regulated: KAAG1*, DNAJB7/HSP40*, TMEM151B, ZBTB32# p &lt; 0.001 ; down-regulated: CPB1*, FOS, TPPP3#, CLEC3B#p &lt; 0.001). Top canonical pathways altered in benign ER+BC included MAPK, PI3K, and acute phase response pathways (p&lt;0.05). GSEA of the entire gene set (N=15,223; ranked in order of 144 DE genes) identified 72 altered pathways (P &lt; 0.005); those with the highest normalization enrichment scores (NES) (&gt; 0.4) functionally grouped as immune function-related (T cell function and antigen presentation). Depleted pathways with NES &gt; 0.4, (N=6) functionally grouped into proteasome-related, fatty acid biosynthesis and mitochondrial energy metabolism. Among the non-coding DE gene set, notably, the entire DE antisense RNA gene set (N=51 transcripts) was up-regulated in benign ER+ BC compared to benign ER-BC (P&lt; 0.001) with a subset (N=11) showing marked up-regulation (&gt; 4 log2FC). Among the DE antisense RNAs, 70% have reported roles in carcinogenesis or BC progression (e.g. KRT7-AS, NAV2-AS2, CCDC144NL-AS1, RP11-66B24.4, HSA-MIR4454). Conclusion: The benign breast transcriptome differs between postmenopausal women with ER+ vs ER- BC, with distinctive coding and non-coding RNA signatures. In postmenopausal women with ER+ BC, benign breast expresses a unique antisense RNA set and is enriched in genes implicated in BC development or progression. These data provide insight into at-risk benign breast and facilitate identification of potential biomarkers of carcinogenesis. Citation Format: Carter JM, Nair AA, Davila JI, Heinzen EP, Hoskin TL, Winham SJ, Radisky DC, Visscher DW, Degnim AC. A unique coding and non-coding benign breast transcriptome in post-menopausal ER+ breast cancer [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 P3-08-10.

Background: Two-staged implant-based reconstruction (IBR) is the most common breast reconstructive modality. Recently, technological and surgical advances have encouraged surgeons to revisit prepectoral IBR. Data comparing prepectoral against subpectoral IBR in women under the age of 40 are lacking. Methods: Retrospective chart review of patients under the age of 40 years old, who underwent immediate 2-staged IBR at our institution, was performed. Patient’s demographics, clinical characteristics, operative details, and early surgical outcomes of prepectoral and subpectoral reconstruction were compared. Data with values of P < 0.05 were considered statistically significant. Results: Between 2012 and 2016, 100 patients (187 breasts) who underwent prepectoral and 69 patients (124 breasts) who underwent subpectoral IBR were included. Median follow-up was 17.9 and 17.5 months in the prepectoral and subpectoral groups, respectively. Total number of complications including both stages of reconstruction was 20 (10.7%) and 19 (15.3%) in the prepectoral and subpectoral groups, respectively (P = 0.227). Specific complications, including hematoma, seroma, skin flap necrosis, wound dehiscence, and breast infections, were not significantly different among groups. Ten (5.4%) devices, including implants and tissue expander, required explantation in the prepectoral group and 8 (6.5%) in the subpectoral group (P = 0.683). Explantation was most commonly due to infection (n = 14), and all of them occurred during the first stage (P < 0.001). Conclusions: Early complications and implant explantation rates are comparable among prepectoral and subpectoral breast reconstruction in women under 40 years old. Based on these results, we believe that prepectoral IBR is a safe, reliable, and promising reconstructive option.

11559 Background: Paraneoplastic neurologic disease (PND) is an aberrant immune-mediated response against the nervous system triggered by occult malignancy. Given the rarity of this disease, a paucity of data exists describing breast cancer (BC)-related PND; thus, we sought to further examine this specific population of patients. Methods: Using an institution specific clinical search tool, we identified all patients at our institution from 1997-2016 with a diagnosis of BC-related PND, verified by an international expert. Retrospective review was used to record tumor and treatment factors associated with PND and BC. A descriptive analysis was performed of the characteristics associated with BC-related PND. Results: BC-related PND was diagnosed in 58 female patients. Average age at BC diagnosis was 54.0±10.3 years. Most had invasive cancer of Stage II or higher [Stage 0 (10.3%), I (10.3%), II (44.8%), III (13.8%), IV (6.9%), unknown (13.8%)]. Most patients were hormone receptor (HR)+ (63.8%), while 4 (6.9%) were HR-, and 17 (29.3%) were unknown. Of the 34 patients where Her2/neu (Her2) data was available, 7 (20.6%) were Her2+ and 27 (79.4%) were Her2-. The majority (62.1%) had neurological symptoms prior to BC, 5.2% were concurrent, and 32.7% developed PND after initial BC. The interquartile range for time from PND to BC diagnosis was -8.5 to 10.5 months. Of the 58 patients, 29 (50%) had autoantibodies detected: Purkinje Cell Cytoplasmic Autoantibody Type-1 (PCA-1[anti-Yo]) (10), amphiphysin-IgG (n = 9), Anti-Neuronal Nuclear Autoantibody Type-2 (ANNA-2[anti-Ri]) (n = 5), and other (n = 5). The most common syndromes were cerebellar ataxia (n = 14), myelopathy (n = 11), and myopathy (n = 11). Immunotherapy was used for PND in 53 of 58 (91.4%) patients; benefit from immunotherapy was assessed as complete 0 (0%), robust 11 (20.8%), mild to moderate 28 (52.8%), none 9 (17%), and indeterminate 5 (9.4%). Conclusions: BC-related PND is rare, with only 58 cases over 20 years at a large academic institution. Paraneoplastic symptoms often present prior to BC diagnosis and BC characteristics are typical of the general population. Over 75% of patients benefit from immunotherapy. These data may provide helpful information to providers treating this population of patients.