High-risk Premenopausal Women Research Articles

Adjuvant therapy for HR+/HER2– early-stage breast cancer (EBC): Changes in practice patterns among community oncologists in Brazil.

e18820 Background: The management of HR+/HER2– early breast cancer (EBC) has changed significantly, with more patients are being classified as low-risk and spared adjuvant chemotherapy (CT) and de-escalating endocrine therapy (ET); on the other hand, other patient groups have experienced treatment escalation, such as high-risk premenopausal women. This study aims to assess practice patterns and real-world data for patients with HR+/HER2- EBC treated in Brazil's largest network of community oncology practices. Methods: Retrospective real-world study from the Oncoclínicas with longitudinal EHR data merged in a platform where structured variables were integrated with unstructured elements from physician notes using technology-based abstraction and expert human curators that follow mCODE standards and predefined ontology. Index data was EBC diagnosis date, and we restricted the study to patients diagnosed between Jan 2017 and Jun 2022, with comparative analysis of adjuvant CT/ET practice trends in 2017-2019 versus 2020-2022. Results: From over 30k patients with breast cancer in Oncoclínicas Database, 19k had HR+/HER2– EBC, 12k were treated in high-volume clinics, and 2,988 were selected for the study due to data accessibility in the pre-specific observation period. Stage distribution: I in 45%, II in 37%, and III in 18%. Pre-menopausal status at diagnosis: 22%. Less than 5% of the study population had access to breast cancer prognostic risk scores. The disease-free survival rate in the entire cohort was 87% (CI95% 86-89) at 2 years and 74% (CI95% 70-78) at 5 years. Overall, we observed a decrease in adjuvant CT use from 51% during 2017-2019 to 41% during 2020-2022. In pre- menopausal patients, CT use reduced from 68% in stage I during 2017-2019 to 41% during 2020-2022; for stage II or III, the reduction was less noticeable (83% in 2017-2019 to 75% in 2020-2022). In post-menopausal patients, CT use reduced from 36% in stage I during 2017-2019 to 18% during 2020-2022; while it remained stable in stage II or III (55% in 2017-2019 versus 52% in 2020-2022). Concerning ET, there were no evident changes in treatment practice. In pre-menopausal, tamoxifen alone was the first option in 90% with stage I and in 64% with stage II or III, with the stability over the years. In post-menopausal, aromatase inhibitors were the first option in 61% with stage I, 70% with stage II and 77% with stage III, with the stability over the years. Conclusions: We perceived a reduction in chemotherapy use in HR+/HER2– stage I breast cancer over time. Reasons for treatment selection must be investigated further, including other prognostic factors, but might be related to greater physician confidence in avoiding CT and access to genomic signatures. There were no significant changes in patterns of adjuvant ET, with most pre- and postmenopausal patients receiving tamoxifen alone and an aromatase inhibitor respectively.

Neither Tumor-Infiltrating Lymphocytes nor Cytotoxic T Cells Predict Enhanced Benefit from Chemotherapy in the DBCG77B Phase III Clinical Trial.

Simple SummaryApart from the direct killing of cancer cells, cyclophosphamide-based chemotherapy has been shown to induce an antitumor immune response, and is being used in combination with immunotherapies in cancer care. We assessed the interaction of chemotherapy with immune biomarkers expressed on primary tumor tissue from a randomized phase III clinical trial, and confirmed that the presence of tumor-infiltrating lymphocytes is linked to improved survival in premenopausal women with high-risk breast cancer, regardless of their treatment allocation. However, immune biomarkers including tumor-infiltrating lymphocytes do not predict extra benefit from cyclophosphamide chemotherapy. This finding applies across the major molecular subgroups, including non-luminal and basal breast cancers that tend to be more immunogenic, and are often considered the most suitable subsets for receiving immunotherapy.Recent studies have shown that immune infiltrates in the tumor microenvironment play a role in response to therapy, with some suggesting that patients with immunogenic tumors may receive increased benefit from chemotherapies. We evaluated this hypothesis in early breast cancer by testing the interaction between immune biomarkers and chemotherapy using materials from DBCG77B, a phase III clinical trial where high-risk premenopausal women were randomized to receive chemotherapy or no chemotherapy. Tissue microarrays were evaluated for tumor-infiltrating lymphocytes (TILs) assessed morphologically on hematoxylin and eosin-stained slides, and by immunohistochemistry for CD8, FOXP3, LAG-3, PD-1 and PD-L1. Following REMARK reporting guidelines, data analyses were performed according to a prespecified statistical plan, using 10-year invasive disease-free survival as the endpoint. Differences in survival probabilities between biomarker groups were evaluated by Kaplan–Meier and Cox proportional hazard ratio analyses and prediction for treatment benefit by an interaction test. Our results showed that stromal TILs were associated with an improved prognosis (HR = 0.93; p-value = 0.03), consistent with previous studies. However, none of the immune biomarkers predicted benefit from chemotherapy in the full study set nor within major breast cancer subtypes. Our study indicates that primary tumors with higher immune infiltration do not derive extra benefit from cyclophosphamide-based cytotoxic chemotherapy.

Abstract 2206: Pilot phase I clinical trial of RANKL inhibition and breast tissue gene expression in high-risk premenopausal women with dense breasts

Abstract Background: Identifying pathways that can be targeted to reduce breast density and breast cancer incidence is an unmet need, especially in premenopausal women. Receptor activator of nuclear factor-κB ligand (RANKL) signaling mediates the major proliferative response of mammary epithelium to progesterone and is positively associated with breast density but there are no clinical trial data on the impact of RANKL inhibition on breast tissue markers in women with dense breasts. We, therefore, determined the impact of RANKL inhibition on breast tissue gene expression in high-risk premenopausal women with dense breasts. Study design: Pilot phase I clinical trial of 9 healthy high-risk pre-menopausal women (≥35 years of age) with dense breasts performed at Washington University School of Medicine, St. Louis, MO from July 2018-December 2018. Fifty-five percent of participants had a positive family history of breast cancer in a first degree relative. Intervention: Participants were given a single dose of subcutaneous RANKL antibody, denosumab (60mg), at baseline (Day 1). On the same day, prior to the denosumab injection, participants had an ultrasound guided core needle biopsy and a blood draw. All study participants returned for repeat core needle biopsy and blood draw after 60 days (Day 60). Outcome Measures: Changes in breast tissue gene expression between Day 1 and Day 60 with a focus on immune/inflammatory and hormone markers. Gene expression was profiled using NanoString nCounter platform. We performed pathway enrichment analysis and used Cell Maker Enrichment library from EnrichR to identify differentially expressed genes. Genes with a p-value<0.05 on differential expression analysis were considered statistically significant for utilization in pathway enrichment analysis. Libraries utilized for pathway enrichment using the EnrichR platform included the ChEA transcription factor library, GO Biological Process library, and Cellular Components library. Results: Macrophage markers, including CCR5, CD86, CD300A, and CD84 and targets downstream of IRF8: (p-value =4.73e-7), neutrophil degranulation/neutrophil activation in the immune response (p-value=6.0e-21), and the secretory granule membrane pathways (p-value=2.4e-14) were downregulated in breast tissues on Day 60. Pathways involved in progesterone metabolism (p-value=0.0003), estrogen response (p-value=0.0014) and fatty acid metabolism, (p-value=0.0001) were significantly upregulated on Day 60. Specifically, genes involved in steroid hormone metabolism such as DHRS2 (p-value=3.13e-05), and AKR1B15: p-value=4.12e-05) were upregulated on Day 60. Conclusions: A single 60 mg dose of subcutaneous denosumab injection was associated with alterations in pathways involved in hormone, immune and inflammatory signaling in the breast tissues of healthy premenopausal women with dense breasts. Citation Format: Michael Waters, Kay Jayachandran, Debbie Bennet, Jin Zhang, Katherine Weilbaecher, Ian S. Hagemann, Graham A. Colditz, Catherine M. Appleton, Adetunji T. Toriola. Pilot phase I clinical trial of RANKL inhibition and breast tissue gene expression in high-risk premenopausal women with dense breasts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2206.

Efficacy and safety of endocrine therapy for breast-cancer prevention in high-risk premenopausal or postmenopausal women: a Bayesian network meta-analysis of nine randomized controlled trials.

Findings in this work might provide certain guidance for current clinical work. This study aimed to evaluate the efficacy and safety of these drugs based on the Bayesian network meta-analysis. Two researchers systematically and comprehensively searched PubMed, Embase, and the central databases of the Cochrane Library from inception to September 15, 2020. The number of specific events and sample size were extracted from each of the included studies. This Bayesian theory-based network meta-analysis included indirect comparisons and mixed treatment analysis. Indirect comparisons compare the efficacy of at least three interventions simultaneously and are mostly used when there are few direct comparison studies. In addition, indirect comparisons are conducted on the basis of direct comparisons through mixed treatment analysis, which can thus improve the accuracy of analysis. A total of nine randomized controlled trials involving 60,732 participants were included. As a result, compared with placebo in high-risk pre- or postmenopausal women, endocrine therapy (ET) decreased the risks of total breast cancer (TBC, odds ratio [OR] 0.69, 95% confidence interval [CI] 0.56-0.85), invasive breast cancer (IBC, OR 0.69, 95% CI 0.53-0.89), estrogen receptor-positive breast cancer (ER+BC) (OR 0.49, 95% CI 0.38-0.64), and ductal carcinoma in situ (OR 0.74, 95% CI 0.56-0.98), but increased the risks of pulmonary embolism (OR 1.33, 95% CI 1.05-1.69), total venous thrombosis (OR 1.75, 95% CI 1.28-2.38), and endometrial carcinoma (EC, OR 1.84, 95% CI 1.17-2.88). In further network stratification analyses, anastrozole, exemestane, and tamoxifen were found to decrease the risks of TBC, IBC, and ER + BC relative to placebo. Similarly, raloxifene decreased the risk of IBC (OR 0.65, 95% CI 0.48-0.85), while tamoxifen increased the risk of EC (OR 2.42, 95% CI 1.10-7.35). To sum up, ET decreased the risks of TBC, IBC, ER + BC, and ductal carcinoma in situ, while increasing the risks of pulmonary embolism, total venous thrombosis, and EC in high-risk pre- or postmenopausal women. Meanwhile, anastrozole, exemestane, and tamoxifen possibly exerted potential protective effects on TBC, IBC and ER + BC. Typically, raloxifene might be effective on IBC, while tamoxifen might increase the risk of EC. Therefore, clinicians should fully weigh the benefits and risks of ET to develop a rational individualized treatment.

Randomized Double-Blind Placebo-Controlled Biomarker Modulation Study of Vitamin D Supplementation in Premenopausal Women at High Risk for Breast Cancer (SWOG S0812).

Observational studies have reported an inverse association between vitamin D intake and breast cancer risk. We examined whether vitamin D supplementation in high-risk premenopausal women reduces mammographic density (MD), an established breast cancer risk factor. We conducted a multicenter randomized double-blind placebo-controlled trial in premenopausal women at high risk for breast cancer [5-year risk ≥ 1.67%, lifetime risk ≥ 20%, lobular carcinoma in situ, prior stage 0-II breast cancer, hereditary breast cancer syndrome, or high MD (heterogeneously/extremely dense)], with a baseline serum 25-hydroxyvitamin D [25(OH)D] ≤ 32 ng/mL. Participants were randomized to 12 months of vitamin D3 20,000 IU/week or matching placebo. The primary endpoint was change in MD from baseline to 12 months using the Cumulus technique. Secondary endpoints included serial blood biomarkers [25(OH)D, 1,25-dihydroxyvitamin D (1,25(OH)D), insulin-like growth factor (IGF)-1, IGF-binding protein-3] and MD change at 24 months. Among 208 women randomized, median age was 44.6 years, 84% were white, 33% had baseline 25(OH)D < 20 ng/mL, and 78% had high baseline MD. Comparing the active and placebo groups at 12 months, MD changes were small and did not significantly differ. Mean MD changes at 12 and 24 months were -0.3% and -1.2%, respectively, in the active arm and +1.5% and +1.6% with placebo (P > 0.05). We observed a mean change in serum 25(OH)D of +18.9 versus +2.8 ng/mL (P < 0.01) and IGF-1 of -9.8 versus -1.8 ng/mL (P = 0.28), respectively. At 12 months, MD was positively correlated with serum IGF-1 and IGF-1/IGFBP-3 (P < 0.01). This trial does not support the use of vitamin D supplementation for breast cancer risk reduction.

What Happens After Menopause? (WHAM): protocol for a prospective, multicentre, age-matched cohort trial of risk-reducing bilateral salpingo-oophorectomy in high-risk premenopausal women

IntroductionWomen at high inherited risk of ovarian cancer are advised to undergo risk-reducing bilateral salpingo-oophorectomy (RRBSO) at age 40–45 years or when their families are complete. Most women are premenopausal...

Increased breast tissue receptor activator of nuclear factor-κB ligand (RANKL) gene expression is associated with higher mammographic density in premenopausal women.

Increased mammographic breast density is associated with a 4–6-fold increased risk of breast cancer, yet lifestyle factors that can reduce dense breasts are yet to be identified, and viable prevention strategies to reduce breast density-associated breast cancer development are yet to be developed. We investigated the associations of breast tissue receptor activator of nuclear factor-κB (RANK) pathway gene expression with mammographic density in 48 premenopausal women, with no previous history of cancer. Gene expression levels were measured in total RNA isolated from formalin-fixed paraffin-embedded breast tissue samples, using the NanoString nCounter platform. Mammographic density was classified based on the American College of Radiology Breast Imaging Reporting and Data (BI-RADS). Linear regression was used to evaluate associations between gene expression and mammographic density. The mean age of participants was 44.4 years. Women with higher breast tissue RANKL (TNFSF11) (p-value = 0.0076), and TNF (p-value = 0.007) gene expression had higher mammographic density. Our finding provides mechanistic support for a breast cancer chemoprevention trial with a RANKL inhibitor among high-risk premenopausal women with dense breasts.

Patient and medical barriers preclude uptake of tamoxifen preventative therapy in women with a strong family history

AimsTo assess the eligibility, uptake and impediments to tamoxifen use in high-risk women attending a risk management clinic due to family history. Patients and methodsAll patients with a germline mutation in a cancer predisposing gene or at high genetic risk (based on family history) attending a Breast and Ovarian cancer risk management clinic from February 2014 to May 2015 received both verbal and written evidence-based information on preventive therapy and were recommended to consider endocrine prevention if not contraindicated. Endocrine therapy initiation, use and cessation were captured. Patient eligibility was analysed and reasons for declining, ceasing or contraindications for medication use were recorded. ResultsDuring the study period, 237 women were seen over 305 consultations for breast surveillance and preventative therapy discussion. They comprised 38 BRCA1 and 42 BRCA2 mutation carriers, 4 with Peutz-Jegher syndrome, 153 with a strong family history. Their median age was 39.4 years. Endocrine preventative was considered and discussed with all but 19 women. Of the remaining 218, 34 chose bilateral prophylactic mastectomy, while endocrine preventative was not recommended in 50 women due to contraindications and 25 women declined treatment due to their intention to fall pregnant. In 118 patients who remained eligible, 18.6% (22) tried prevention and 9.4% (14) remained on therapy. ConclusionsPhysician-reluctance is not a dominant reason for poor uptake of endocrine prevention even by high-risk premenopausal women in a specialised risk management clinic. Many women are not eligible, and most elect for alternative options.

Exercise-Induced Dose-Response Alterations in Adiponectin and Leptin Levels Are Dependent on Body Fat Changes in Women at Risk for Breast Cancer.

Dysregulation of adipokines, such as adiponectin and leptin, is associated with a variety of chronic diseases, including cancer. Physical activity protects against breast cancer and one of the mechanisms which may underlie this association is exercise-induced changes in adipokine levels. The WISER Sister Trial was a three-armed randomized controlled trial in premenopausal women (n = 137) with an elevated risk for breast cancer. A 5-menstrual-cycle-long dosed aerobic exercise intervention compared low-dose exercise (150 min/wk; n = 44) or high-dose exercise (300 min/wk; n = 48) with a control group asked to maintain usual activity levels (n = 45). Exercise intensity progressed to and was maintained at 70% to 80% of age predicted heart rate max. Body composition and adipokine levels were measured at baseline and follow-up. We observed significant linear trends for increased fitness capacity (Δ%: -2.0% control, 10.1% low dose, 13.1% high dose), decreased fat tissue-to-total tissue mass (Δ%: 0.7% control, -2.9% low dose, -3.7% high dose), increased body fat adjusted adiponectin (Δ%: -0.6% control, 0.6% low dose, 0.9% high dose), and decreased body fat adjusted leptin (Δ%: 0.7% control, -8.2% low dose, -10.2% high dose). In this randomized clinical trial of premenopausal women at risk for breast cancer, we demonstrate a dose-response effect of exercise on adiponectin and leptin and that dose response is dependent on changes in body fat. Improved adipokine levels, achieved by aerobic exercise training-induced decreases in body fat, may decrease breast cancer risk for high-risk premenopausal women. Cancer Epidemiol Biomarkers Prev; 25(8); 1195-200. ©2016 AACR.

Abstract 1789: Estrogen receptor alpha and beta gene expression in benign breast tissue of high risk premenopausal women treated with the SERM acolbifene

Abstract Background We conducted a pilot study to assess the feasibility of a 4th generation selective estrogen receptor modulator (SERM) acolbifene as a breast cancer prevention agent in premenopausal women at high risk for development of breast cancer [Cancer Prev Res 8, December 2015]. Favorable modulation of biomarkers was observed in benign breast tissue acquired before and after intervention. This included reduction in proliferation (Ki-67 immunocytochemistry) and decreases in expression of genes that code for ERα, pS2, and PgR. While ERα is over-expressed and stimulatory in breast carcinogenesis, ERβ which would be inhibitory is down-regulated. Thus, a change in the ratio of gene expression might be most informative. We have now analyzed gene expression of ERβ so as to examine whether it, or the ratio of ERα to ERβ, may contribute to assessment of response. Methods Premenopausal women at elevated risk for breast cancer were screened by random periareolar fine needle aspiration (RPFNA) performed during the follicular phase of the menstrual cycle. Eligible women (n = 25) received 6-8 months of open-label acolbifene (20 mg/d), followed by repeat RPFNA. Reverse-transcription quantitative polymerase chain reaction (RT-qPCR) was performed on 16 paired specimens for several genes associated with estrogen activity: estrogen receptor 1 (ESR1) for ERα, trefoil factor 1 (TFF1) for pS2, GREB1 for growth regulation by estrogen in breast cancer 1, PGR for progesterone receptor, and estrogen receptor 2 (ESR2) for ERβ. The quantity of each biomarker transcript was expressed relative to the reference transcript HPRT1. Further normalization by epithelial cell markers (KRT19 for cytokeratin 19 or CDH1 for E-Cadherin) did not materially alter the results. Results Significant changes had been previously shown for transcripts that code for pS2 (15 decreases, 1 increases; p = 0.002), ERα (11↓, 5↑; p = 0.013), and PgR (15↓, 1↑; p = 0.001); plus a suggestion of effect of GREB-1α (10↓, 6↑; p = 0.074). ERβ exhibited a general increase, but this was not statistically significant (5↓, 11↑; p = 0.24). However, the reduction of the ratio of ERα to ERβ was statistically significant (15↓, 1↑; p = 0.002). Conclusions Assessment of the ratio of gene expression for ERα to ERβ, and possibly their protein products by immunocytochemistry, may provide a useful surrogate endpoint biomarker by which to monitor response to SERMs used for breast cancer chemoprevention in premenopausal women. Supported by NO1-CN-35135. Citation Format: Bruce F. Kimler, Teresa A. Phillips, Carol J. Fabian. Estrogen receptor alpha and beta gene expression in benign breast tissue of high risk premenopausal women treated with the SERM acolbifene. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1789.

The Effect of Atorvastatin on Breast Cancer Biomarkers in High-Risk Women.

Statins have the potential to reduce breast cancer incidence and recurrence as shown in both epidemiologic and laboratory studies. The purpose of this study was to evaluate the effect of a lipophilic statin, atorvastatin, on breast cancer biomarkers of risk [mammographic density (MD) and insulin growth factor 1 (IGF-1)] in high-risk premenopausal women.Premenopausal women at increased risk for breast cancer received either 40 mg of atorvastatin or placebo for 1 year. Biomarker assessment was performed prior to initiation and at completion of study medication. MD was determined using both Breast Imaging Reporting and Data System and the visual analogue scale. Serum IGF-1 was determined by ELISA assay at the end of the study.Sixty-three women were enrolled between December 2005 and May 2010. Sixteen (25%) women withdrew. The mean age of participants was 43 (range, 35-50), 100% were white, and the average body mass index (BMI) was 26.4. The statin group demonstrated a significant decrease in cholesterol and low-density lipoprotein (LDL), suggesting compliance with study medication. After accounting for BMI, there was no difference in change in MD between groups. There was a significant increase in serum IGF-1 in the statin group.In this multi-institutional randomized prospective clinical trial of premenopausal women at increased risk for breast cancer, we did not see an effect of atorvastatin on MD. Further investigation of statins may be warranted; however, design of prior trials and potential mechanism of action of the agent need to be considered in the design of future trials. Cancer Prev Res; 9(5); 379-84. ©2016 AACR.

Options in Prophylactic Surgery to Prevent Ovarian Cancer in High-Risk Women: How New Hypotheses of Fallopian Tube Origin Influence Recommendations.

In women at increased risk of developing ovarian cancer, risk-reducing salpingo-oophorectomy is the only intervention that has been shown to decrease mortality from ovarian cancer and is the standard of care for risk reduction. Prophylactic salpingectomy with delayed oophorectomy should be considered in high-risk premenopausal women in the setting of a clinical trial.

Excess Weight Gain Accelerates 1-Methyl-1-Nitrosourea–Induced Mammary Carcinogenesis in a Rat Model of Premenopausal Breast Cancer

In contrast to the null effects generally reported, high-risk premenopausal women (Gail score ≥1.66) enrolled in the Breast Cancer Prevention P-1 Trial were recently reported to be at increased risk for breast cancer when overweight (HR = 1.59) or obese (HR = 1.70). To investigate this clinical observation in a preclinical setting, ovary-intact female rats were intraperitoneally injected with 50 mg/kg 1-methyl-1-nitrosourea at 21 days of age to simulate premenopausal women with increased risk. Two commercially available strains of Sprague-Dawley rat (Taconic Farms) were used, which are dietary resistant (DR) or dietary susceptible (DS) to excess weight gain when fed a purified diet containing 32% kcal from fat, similar to levels consumed by the typical American woman. DS rats were approximately 15.5% heavier than DR rats at study termination and plasma leptin indicated a marked difference in adiposity. DS rats had higher incidence (26% increase), multiplicity (2.5-fold increase), and burden (5.4-fold increase) of mammary carcinomas with a concomitant reduction in cancer latency (16% earlier detection) compared with DR rats (P < 0.001 for all analyses), and displayed a higher proportion of hormone receptor negative tumors compared with DR rats [OR = 1.78; 95% confidence interval (CI), 0.83-3.81]. Circulating levels of several breast cancer-risk factors, including leptin, adiponectin:leptin ratio, insulin, insulin-like growth factor (IGF)-1, IGF-1:IGF-1 binding protein-3 ratio, and calculated insulin resistance (HOMA-IR) were negatively impacted in DS rats (P < 0.05 for all analyses). These findings support further investigation of the effects of excess weight in high-risk premenopausal women and demonstrate a useful preclinical model for rapid evaluation of mechanistic hypotheses.

RSF1 and Not Cyclin D1 Gene Amplification May Predict Lack of Benefit from Adjuvant Tamoxifen in High-Risk Pre-Menopausal Women in the MA.12 Randomized Clinical Trial

Most women with estrogen receptor expressing breast cancers receiving anti-estrogens such as tamoxifen may not need or benefit from them. Besides the estrogen receptor, there are no predictive biomarkers to help select breast cancer patients for tamoxifen treatment. CCND1 (cyclin D1) gene amplification is a putative candidate tamoxifen predictive biomarker. The RSF1 (remodeling and spacing factor 1) gene is frequently co-amplified with CCND1 on chromosome 11q. We validated the predictive value of these biomarkers in the MA.12 randomized study of adjuvant tamoxifen vs. placebo in high-risk premenopausal early breast cancer. Premenopausal women with node-positive/high-risk node-negative early breast cancer received standard adjuvant chemotherapy and then were randomized to tamoxifen (20 mg/day) or placebo for 5 yrs. Overall survival (OS) and relapse-free survival (RFS) were evaluated. Fluorescent in-situ hybridization was performed on a tissue microarray of 495 breast tumors (74% of patients) to measure CCND1 and RSF1 copy number. A multivariate Cox model to obtain hazard ratios (HR) adjusting for clinico-pathologic factors was used to assess the effect of these biomarkers on Os and RFS. 672 women were followed for a median of 8.4 years. We were able to measure the DNA copy number of CCND1 in 442 patients and RSF1 in 413 patients. CCND1 gene amplification was observed in 8.7% and RSF1 in 6.8% of these patients, preferentially in estrogen receptor-positive breast cancers. No statistically significant interaction with treatment was observed for either CCND1 or RSF1 amplification, although patients with high RSF1 copy number did not show benefit from adjuvant tamoxifen (HR = 1.11, interaction p = 0.09). Unlike CCND1 amplification, RSF1 amplification may predict for outcome in high-risk premenopausal breast cancer patients treated with adjuvant tamoxifen.

Patterns of Nonmasslike Enhancement at Screening Breast MR Imaging of High-Risk Premenopausal Women

Current U.S. recommendations for breast cancer screening of women with at least a 20%-25% lifetime risk of developing breast cancer include contrast material-enhanced magnetic resonance (MR) imaging of the breasts. The cancer detection rate in high-risk women undergoing screening MR imaging is approximately 10 times higher than that in normal-risk women undergoing screening mammography. Many of these high-risk women commence MR imaging screening while they are premenopausal, when the breasts are most influenced by cyclical hormonal changes. Healthy premenopausal breast tissue enhances in a cyclical and variable manner. This enhancement is described as background enhancement. Typically, enhancement of normal breast tissue occurs in a symmetric and diffuse pattern, and there is little diagnostic difficulty in classifying it as normal background parenchymal enhancement. However, sometimes the pattern is more focal, asymmetric, or regional. It may then be described as nonmasslike enhancement, an observation associated with both benign and malignant breast pathologic conditions. A review of the morphologic features and internal enhancement patterns in normal but nondiffuse background enhancement and abnormal nonmasslike enhancement in high-risk premenopausal women can help improve interpretive specificity and decrease false-positive interpretations. MR imaging pitfalls and interpretation strategies for localized background enhancement and pathologic nonmasslike enhancement in this high-risk population are highlighted. In evaluating nonmasslike enhancement, the use of the Breast Imaging Reporting and Data System (BI-RADS) lexicon to perform careful analysis of morphologic features, along with an understanding of the role and limitations of kinetic information, will help balance early breast cancer detection against false-positive interpretation.

Abstract 3601: Excess weight gain accelerates 1-methyl-1-nitrosourea (MNU)-induced mammary carcinogenesis in ovary-intact Sprague Dawley (Levin) rats fed 31.8% dietary calories from fat.

Abstract Though population data for breast cancer indicate that adult weight gain is associated with increased risk in postmenopausal women, the generally accepted lack of effect of obesity on premenopausal breast cancer risk has recently been challenged (Cecchini et al Cancer Prev Res; 5(4); 583-92, 2012). In that publication from the National Surgical Adjuvant Breast and Bowel Project (NSABP), BMI &amp;gt;30 was significantly associated with increased risk of invasive breast cancer in high-risk premenopausal women (HR= 1.70). As the effects of excess weight gain on breast carcinogenesis in women prior to menopause remain poorly understood, the Levin strains of outbred Sprague Dawley rats that are dietary obesity sensitive (DS) or resistant (DR) were used to investigate this issue. The Levin rats, which are available from Taconic Farms, were selectively outbred for &amp;gt;20 generations for resistance (DR) or susceptibility (DS) to weight gain when fed a purified diet of similar fat content as the typical diet consumed by women in the United States (31.8% fat by kcal). DS rats gain more weight than DR rats and develop metabolic syndrome after several weeks, including dyslipidemia and insulin resistance. Ovary-intact DS (n=101) and DR (n=103) rats were injected with 50 mg/kg of 1-methyl-1-nitrosourea (MNU) at 21-24 days of age, and began ad libitum feeding of the purified diet the following day. The experiment was terminated 9 weeks post-carcinogen at which point DS rats were approximately 17% heavier than their DR counterparts (220.3 ± 22.1 g vs. 187.9 ± 14.6 g, p&amp;lt;0.0001). The incidence, multiplicity, and tumor burden per rat of histopathologically confirmed palpable mammary carcinomas were significantly higher in the DS versus DR rats (incidence 93.07% vs. 64.08%, p&amp;lt;0.05; multiplicity, 4.17 tumors vs. 1.82 tumors, p&amp;lt;0.01; tumor burden, 7.42 g vs. 1.93 g, p&amp;lt;0.0001) and cancer latency was reduced (37.9 ± 9.0 days vs. 44.0 ± 6.7 days, p&amp;lt;0.0001). Molecular subtypes of tumors in DS and DR are currently being evaluated. These results suggest that the Levin model of DS and DR rats represents a valuable preclinical tool for further investigating how excessive energy intake and weight gain with accompanying metabolic syndrome affect the carcinogenic process in the breast. Supported by PHS grant CA52626 from the National Cancer Institute. Citation Format: Shawna B. Matthews, John N. McGinley, Zongjian Zhu, Weiqin Jiang, Elizabeth Neil, Angie Neil, Henry J. Thompson. Excess weight gain accelerates 1-methyl-1-nitrosourea (MNU)-induced mammary carcinogenesis in ovary-intact Sprague Dawley (Levin) rats fed 31.8% dietary calories from fat. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3601. doi:10.1158/1538-7445.AM2013-3601

Abstract P1-03-01: Evidence for the Warburg effect in mammary atypia from high-risk African American women.

Abstract Background: Aggressive cancers are known to consume glucose avidly and produce lactic acid (rather than fully metabolize glucose via the Tricarboxylic Acid (TCA) cycle). This shift toward lactate production, even in the presence of adequate oxygen, is termed the Warburg effect. The Warburg effect is thought to be a late event in breast cancer, however, our studies in high-risk women provide evidence that the Warburg effect occurs during cancer initiation. This is an important observation as glucose-signaling can be readily targeted for breast cancer prevention with minimal toxicity. Here we investigated the role of the Warburg effect in breast cancer initiation in young high-risk women. Methods and Results: Similar to fluorodeoxyglucose, 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG) is a fluorescent glucose analog that can be used to track glucose uptake and glycolysis. 2-NBDG spectroscopy provides a means to track glucose metabolism in live mammary epithelial cells from high-risk women. We used 2-NBDG spectroscopy to measure glucose uptake in ER− breast cancer and live atypical mammary epithelial cells from high-risk premenopausal women. We observe that both triple-negative breast cancer and a subset of atypia exhibits accumulation of 2-NBDG. There is growing recognition that phosphoprotein signaling networks (rather than single genes) play a key role in breast cancer initiation and progression. Our team used Reverse Phase Proteomic Microarray (RPPM) profiling to test for activation of phosphoprotein signaling networks in atypical RPFNA cytology from high-risk premenopausal women in our cohort. RPFNA were obtained from two independent sets of 39 and 38 high-risk premenopausal women; 45% of these women were African American. The signaling network most highly expressed in precancerous cells contained activated signaling proteins associated with the Warburg effect (AKT/mTOR/PI3K), insulin signaling (pACC, IRS1) and epithelial to mesenchymal transition (EMT) IL6/Stat3/vimentin. Conclusions: This is the first evidence that abnormal glucose uptake and the Warburg effect occurs during breast cancer initiation in high-risk premenopausal women. These studies demonstrate our ability to identify abnormal glucose and activated signaling networks associated with the Warbug effect in atypical mammary cells from high-risk women. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P1-03-01.

Abstract P1-09-02: Pilot study of a 1-year intervention of high-dose vitamin D in women at high risk for breast cancer.

Abstract Background: Selective estrogen receptor modulators (SERMs) have been shown to decrease breast cancer incidence among high-risk women, but uptake for prevention has been poor. Observational studies have demonstrated that serum 25-hydroxyvitamin D (25-OHD) is inversely related to breast cancer risk, such that levels &amp;gt;40 ng/ml are associated with about a 40% reduction in breast cancer risk compared to women who are vitamin D deficient (25-OHD &amp;lt;20 ng/ml). Uncertainty remains about whether vitamin D supplementation will reduce breast cancer risk, the optimal dose of vitamin D, and the target level of serum 25-OHD. We examined the safety of high dose vitamin D and the effects on serum 25-OHD in women at high risk for breast cancer. Methods: Forty high-risk women (defined as a 5-year Gail risk ³1.67%, lobular or ductal carcinoma in situ [LCIS/DCIS], BRCA1/BRCA2 mutation carrier, or stage I/II invasive breast cancer in remission for &amp;gt;5 years) were assigned to a 1-year intervention of vitamin D3 20,000 or 30,000 IU weekly. Other eligibility criteria included baseline mammographic density (MD) ≥25%, serum 25-OHD ≤32 ng/ml, no current SERM use and no history of kidney stones. Women underwent a digital mammogram at baseline and 12 months, and serial blood draws every 3 months. In addition, random core breast biopsies were conducted in premenopausal women, whereas postmenopausal women underwent a breast MRI at baseline and 12 months. Participants were monitored for toxicity, particularly hypercalcemia and hypercalciuria, every 3 months. The primary objective is to determine the safety and feasibility of high-dose vitamin D in this study population. Secondary objectives are to determine changes in breast density and blood-based biomarkers (25-OHD, 1,25(OH)D, PTH, IGF-I, IGFBP-3). Serum 25-OHD was measured by Diasorin radioimmunoassay. Results: From November 2007 to January 2011, 292 women were screened and 142 were ineligible. Main reasons for ineligibility (%) included 25-OHD &amp;gt;32 ng/ml (27), opted for SERM (23), prior kidney stones (11), and MD &amp;lt;25% (9). Of the 40 enrolled participants: median age 50 years (range, 37–73); pre/postmenopausal: 20/20; white/hispanic/black/asian: 19/14/6/1; median body mass index 26.6 kg/m2 (20–39.6); elevated Gail risk/LCIS/DCIS/stage I or II breast cancer: 20/10/8/2; mean baseline serum 25-OHD 20.2 ng/ml (9–31). Currently, 1 participant is on-study, 31 completed the intervention, 6 were lost to follow-up, 1 withdrew due to hypercalciuria (spot urine Ca/Cr &amp;gt;0.37) and 1 withdrew due to dyspepsia. Mean serum 25-OHD rose to 47 ng/ml at 3 months, 49.1 ng/ml at 6 months, and 53.7 ng/ml (range, 26–77) at 12 months. No significant hypercalcemia (serum Ca &amp;gt;10.5 mg/dl) occurred at either dose level. Imaging and biomarker analyses are ongoing. Discussion: We have demonstrated that a 1-year intervention of high-dose vitamin D3 is well tolerated and can increase serum 25-OHD above a target level of 40 ng/ml. This preliminary data has informed an ongoing phase IIb randomized placebo-controlled trial (SWOG 0812) of high-dose vitamin D in 200 high-risk premenopausal women and highlights the importance of early phase breast cancer chemoprevention trials with intermediate biomarker endpoints to test novel agents. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P1-09-02.

Reduction in Ki-67 in benign breast tissue of high-risk premenopausal women with the SERM acolbifene.

520 Background: Selective Estrogen Receptor Modulators (SERMs) are approved for reduction of risk for breast cancer; however, uptake and use is limited. We conducted a pilot study of a 4th generation SERM to determine tolerability and effect on tissue biomarkers in healthy women at high risk for development of breast cancer. Methods: Premenopausal women at elevated risk for breast cancer were screened by random periareolar fine needle aspiration (RPFNA) performed during the follicular phase of the menstrual cycle. Women were eligible if breast epithelial cells exhibited evidence of cytologic hyperplasia with or without atypia, as well as Ki-67 ≥2% by immunocytochemistry. Following 6-8 months of open-label acolbifene (20 mg/d), the RPFNA was repeated. The primary endpoint was modulation of the proportion of cells that expressed Ki-67. Body composition (DEXA), pelvic sonography, mammographic breast density, and serum levels of IGF-1/IGFBP3 and several bioavailable hormones were assessed pre and post intervention. Results: 76 women were screened by RPFNA, with 25 eligible and enrolled in the intervention over a 9 month period. All 25 (7 on oral contraceptives) subjects completed the study, had a second RPFNA, and were evaluable. Median Ki-67 at baseline was 4.6% (range 2.4 – 21.9%) and off study 1.4% (range 0 – 6.6%); median change was a reduction of 3.0% (range -20.2% to +2.8%; decreased in 23, increased in 2) or a relative reduction of 77%. The end-of-study Ki-67 was significantly less than baseline (p&lt;0.001, 2-tailed Wilcoxon test). There were no statistically significant changes in cytomorphology over this short intervention period. There was a marginal effect on breast density (16 decreased; 8 increased; p=0.067). Adverse events were minimal with greatest grade of 3 reported by 2 subjects, grade 2 by 7 subjects, and grade 1 by 11 subjects. No serious adverse event was reported and no subject discontinued the study due to an AE. Conclusions: Based on preliminary evaluation showing favorable modulation of proliferation and minimal adverse events, further investigation of acolbifene, a fourth generation SERM, as a breast cancer chemoprevention agent for premenopausal women appears warranted. Supported by NO1-CN-35135.

Abstract 686: Warburg effect in mammary atypia from high-risk women

Abstract Background: Aggressive cancers are known to consume glucose avidly and produce lactic acid (rather than fully metabolize glucose via the Tricarboxylic Acid (TCA) cycle). This shift toward lactate production, even in the presence of adequate oxygen, is termed the Warburg effect. The Warburg effect is thought to be a late event in breast cancer, however, our studies in high-risk women provide evidence that the Warburg effect occurs during cancer initiation. This is an important observation as glucose-signaling can be readily targeted for breast cancer prevention with minimal toxicity. Here we investigated the role of the Warburg effect in breast cancer initiation in young high-risk women. Methods and Results: Similar to fluorodeoxyglucose, 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG) is a fluorescent glucose analog that can be used to track glucose uptake and glycolysis. 2-NBDG spectroscopy provides a means to track glucose metabolism in live mammary epithelial cells from high-risk women. We used 2-NBDG spectroscopy to measure glucose uptake in ER- breast cancer and live atypical mammary epithelial cells from high-risk premenopausal women. We observe that both triple-negative breast cancer and a subset of atypia exhibits accumulation of 2-NBDG. There is growing recognition that phosphoprotein signaling networks (rather than single genes) play a key role in breast cancer initiation and progression. Our team used Reverse Phase Proteomic Microarray (RPPM) profiling to test for activation of phosphoprotein signaling networks in atypical RPFNA cytology from high-risk premenopausal women in our cohort. RPFNA were obtained from two independent sets of 39 and 38 high-risk premenopausal women; 45% of these women were African American. The signaling network most highly expressed in precancerous cells contained activated signaling proteins associated with the Warburg effect (AKT/mTOR/PI3K), insulin signaling (pACC, IRS1) and epithelial to mesenchymal transition (EMT) IL6/Stat3/vimentin. Conclusions: This is the first evidence that abnormal glucose uptake and the Warburg effect occurs during breast cancer initiation in high-risk premenopausal women. These studies demonstrate our ability to identify abnormal glucose and activated signaling networks associated with the Warbug effect in atypical mammary cells from high-risk women. 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 686. doi:1538-7445.AM2012-686