Subsequent Development Of Breast Cancer Research Articles

Fibroepithelial Neoplasm with Hybrid Features of Benign Phyllodes Tumor, Juvenile Papillomatosis, and Juvenile Fibroadenoma: A Case Report.

Phyllodes tumor is an uncommon breast fibroepithelial neoplasm mainly found in middle-aged patients, presenting a morphologic continuum from benign to malignant. Juvenile papillomatosis represents a rare benign proliferative breast tumor primarily affecting young individuals and carries a potential elevated risk of subsequent breast cancer development. Juvenile fibroadenoma is a well-circumscribed biphasic neoplasm that often occurs in adolescent girls, characterized by a pericanalicular growth pattern with usual-type epithelial hyperplasia and gynaecomastia-like micropapillary proliferation. Herein, we present an unusual example of a 26-year-old woman with a left breast outer lower quadrant palpable mass. Ultrasonography identified a 5.9 cm lobulated hypoechoic solid mass with scattered small cysts. The preoperative biopsy initially diagnosed a fibroepithelial lesion, considering giant cellular fibroadenoma and phyllodes tumor in the differential. Subsequent complete excision revealed areas of benign phyllodes tumor features closely admixed with distinctive elements such as prominent multiple cysts exhibiting apocrine and papillary apocrine metaplasia, duct papillomatosis, and duct stasis characteristic of juvenile papillomatosis, and hyperplastic ductal epithelium with micropapillary projections demonstrating a pericanalicular growth pattern indicative of juvenile fibroadenoma. The diagnosis was conclusively established as a fibroepithelial lesion with combined features of benign phyllodes tumor, juvenile papillomatosis, and juvenile fibroadenoma. Further investigation uncovered a family history of breast cancer. Molecular analysis revealed a pattern of unique and overlapping mutations within these distinct histopathological areas. This unusual presentation with hybrid features within a single tumor is described for the first time in the literature along with the molecular signature of the individual components.

Abstract 283: Fibroblast diversity and immune microenvironment in pseudoangiomatous stromal hyperplasia (PASH): Associations with breast cancer risk

Abstract Benign breast disease (BBD) affects over one million women annually in the United States and is diagnosed in nearly half of all women during their lifetime. Pseudoangiomatous stromal hyperplasia (PASH), a common BBD subtype, is characterized by an overaccumulation of tissue fibroblasts. Current research suggests a pivotal role of the immune microenvironment in progression of BBD to breast cancer, suggesting its composition as a determinant of breast cancer risk. Our study evaluates whether fibroblast variations and immune microenvironment differences in BBD biopsies containing PASH lesions are associated with subsequent breast cancer development. We evaluated BBD biopsies from women who subsequently developed breast cancer (cases, N=5) or remained cancer-free (controls, N=5). Using Opal-based multiplex immunofluorescence (mIF) assays, we mapped expression of fibroblast markers linked to breast cancer associated fibroblasts (CAFs), including vimentin (VIM), smooth muscle actin (ACTA2), fibroblast specific protein-1 (S100A4/FSP1), fibroblast associated protein (FAP), integrin beta-1 (CD29/ITGB1), and platelet derived growth factor receptor beta (PDGFRβ) and markers of key immune cells (T cells: CD4, CD8; dendritic cells: CD11c; B cells: CD20; and macrophages: CD68) along with MKI67 as a proliferation marker. Our results revealed marked differences in fibroblast marker expression and immune cell populations between these groups in the normal lobules in the BBD biopsies. PASH cases showed elevated PDGFRβ and CD29/ITGB1 expression in VIM-positive fibroblasts, markers characteristic of CAF subtypes known to accumulate in more aggressive tumor microenvironments. Conversely, a higher percentage of FSP1+/VIM+ cells, similar to that of a different CAF subtype, was noted in the lobules of PASH controls. This difference in biomarker expression, along with corresponding variations in immune cell counts, suggests a complex interplay between fibroblast composition and immune response that ultimately modulate cancer risk in PASH-containing BBD tissues. Additionally, increased MKI67 expression in PASH cases potentially signals the higher likelihood of cancer development. This work advances our understanding of fibroblast heterogeneity in BBD and its implications for breast cancer risk and progression, and paves the way for innovative strategies to improve clinical outcomes for BBD patients to reduce their incidence of breast cancer. Citation Format: Nicole Cruz-Reyes, Matilde Rossi, Melody Stallings Mann, Bryan McCauley, Tanya Hoskin, Robert Vierkant, Stacey Winham, Amy Degnim, Mark E. Sherman, Derek C. Radisky. Fibroblast diversity and immune microenvironment in pseudoangiomatous stromal hyperplasia (PASH): Associations with breast cancer risk [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 283.

356 Upregulated Genes in Age-Related Lobular Involution Stagnation Represent Potential Biomarkers That Link To Increased Breast Cancer Risk

OBJECTIVES/GOALS: Age-related lobular involution (LI) is a physiological process of breast epithelial regression that occurs primarily during perimenopause (ages 45-55); women in this age range for which the process of LI is delayed, defined as LI stagnation, show significantly increased risk of breast cancer as compared to LI progression patients. METHODS/STUDY POPULATION: The Mayo Clinic Benign Breast Disease (BBD) cohort includes ~1000 women who had multiple sequential benign biopsies. 103 patients were found to have sequential biopsies during the perimenopausal period, of which 10 eventually progressed to breast cancer. These patients were assessed for LI stagnation vs LI progression by quantifying 10 lobules per slide and comparing median acini number and median lobule size between initial and subsequent biopsies from the same patients. RNA was derived from whole tissue sections from the initial biopsies, and profiled using NanoString IO360 and BC360, which were normalized using RUVg methods. Differentially expressed genes associated with LI stagnation were defined as having two-tailed, unpaired p-values less than 0.05. RESULTS/ANTICIPATED RESULTS: Analysis showed subsetting patient sets by time between biopsies improves classification of stagnant vs. progression. Differential gene analysis identified 37 genes associated with LI stagnation and LI progression, and 20 of these genes were found to overlap a set of 128 gnese that were differentially expressed between women who subsequently developed breast cancer vs remained cancer-free. These genes represent potential biomarkers of processes that link LI stagnation and increased breast cancer risk. DISCUSSION/SIGNIFICANCE: In future studies, we intend to study these genes that were shown to be upregulated in LI stagnation for their association with subsequent development of breast cancer in independent cohorts of women with BBD. We will use this knowledge to improve individualized risk assessment, which will help focus surveillance and prevention strategies.

Abstract SY25-02: Double-edged “soil”: Stromal microenvironment in breast cancer development

Abstract SY25-02: Double-edged “soil”: Stromal microenvironment in breast cancer development

Biological features of ductal carcinoma in situ: clinical role and basis for treatment individualization

Ductal carcinoma in situ (DCIS) is an extremely heterogeneous disease in terms of clinical manifestations, morphological changes, and expression of biomarkers, which determine the risk of subsequent development of an invasive breast cancer (BC). Diagnosis and treatment of DCIS prevents the development of invasive tumors (which reduces the potential risk of death from BC); however, the prognostic value of local treatment depends on the biological characteristics of its. The tumor grade, presence of comedonecrosis and expression of estrogen receptors are the key prognostic factors in DCIS for the treatment tactics and prognosis. The clinical symptoms of DCIS are very scarce, the most of tumors is diagnosed by screening mammography; the typical sign of DCIS is malignant microcalcifications or changes of breast architectonic. The sensitivity of mammography, ultrasound and MRI directly depend on the biological characteristics of DCIS. Surgical treatment (breast-conserving surgery or mastectomy) significantly reduces the risk of BC death in women with DCIS G2/G3, and radiotherapy after breast-conserving surgery reduce the risk of local recurrence of non-invasive and invasive BC. The choice of the local treatment (breast-conserving surgery radiation therapy vs mastectomy) depend on such factors as: tumor size, localization, clear margins, and biological characteristics of DCIS. In contrast to invasive cancers, the negative margin in DCIS is more than 2 mm from the tumor. Regional lymph node involvement in DCIS occurs in less than 1% of cases; however, microinvasion is found in analyze the surgical specimen in 15% of patients, which raises the question about regional staging. Risk factors for microinvasion in DCIS are age less than 55 years, tumor lesion size more than 4.0 cm, DCIS grade G3, and tumor palpability. Adjuvant endocrine therapy with tamoxifen significantly reduces the 10-year risk of invasive recurrence by 51%, the risk of contralateral BC by 50% and the risk of death by 40%, but only for hormone-positive DCIS. HER2 overexpression is found in DCIS significantly often (up to 40% of cases) than in invasive BC, HER2+ status correlates with DCIS high grade G3, the presence of comedonecrosis and with increased risk of relapse (both non-invasive and invasive) but is not a reason for anti-HER2 therapy. DCIS low risk and DCIS high risk have not only different morphological characteristics, but also different models of biological behavior, which must be considered in the different choice of treatment tactics.

Modulation of the immune microenvironment of high-risk ductal carcinoma in situ by intralesional pembrolizumab injection

Ductal carcinoma in situ (DCIS) is a risk factor for the subsequent development of invasive breast cancer. High-risk features include age <45 years, size >5 cm, high-grade, palpable mass, hormone receptor negativity, and HER2 positivity. We have previously shown that immune infiltrates are positively associated with these high-risk features, suggesting that manipulating the immune microenvironment in high-risk DCIS could potentially alter disease progression. Patients with high-risk DCIS were enrolled in this 3 × 3 phase 1 dose-escalation pilot study of 2, 4, and 8 mg intralesional injections of the PD-1 immune checkpoint inhibitor, pembrolizumab. Study participants received two intralesional injections, three weeks apart, prior to surgery. Tissue from pre-treatment biopsies and post-treatment surgical resections was analyzed using multiplex immunofluorescence (mIF) staining for various immune cell populations. The intralesional injections were easily administered and well-tolerated. mIF analyses demonstrated significant increases in total T cell and CD8+ T cell percentages in most patients after receiving pembrolizumab, even at the 2 mg dose. T cell expansion was confined primarily to the stroma rather than within DCIS-containing ducts. Neither cleaved caspase 3 (CC3) staining, a marker for apoptosis, nor DCIS volume (as measured by MRI) changed significantly following treatment. Intralesional injection of pembrolizumab is safe and feasible in patients with DCIS. Nearly all patients experienced robust total and CD8+ T cell responses. However, we did not observe evidence of cell death or tumor volume decrease by MRI, suggesting that additional strategies may be needed to elicit stronger anti-tumor immunity.

Gene methylation and cytological atypia in random fine needle aspirates for assessment of breast cancer risk: 10-year follow up in average risk women.

10520 Background: Alterations in DNA methylation occur early in tumorigenesis, and are a potential breast cancer risk biomarker. We previously reported a study where healthy volunteers underwent random fine needle aspiration (rFNA) of the breasts; cumulative methylation index of eight preselected tumor suppressor genes (CMI) was associated with the presence of cytological atypia in rFNA samples. We now report 10-year follow-up of this population, to evaluate whether increased CMI is associated with subsequent breast cancer development. Methods: 380 women, unselected by breast cancer risk, were enrolled. Demographics, breast cancer risk factors, lifetime Gail model risk estimates (Gail-LR), and %breast density were obtained at baseline. rFNA samples were assessed for cytopathology (Masood Score, MS) and CMI. Patients were contacted annually for 10 years to ascertain development of invasive or non-invasive breast cancer. In univariate analysis, log-rank test was used to compare breast cancer incidence rates between individuals with high and low baseline measures (separated by median). Area under the ROC curve was used to evaluate the cancer prediction accuracy. In multivariate analysis, the effect of CMI (after log-transformation to reduce skewness) was further studied using Cox regression model adjusting for confounding baseline variables. Results: 362 women participated in follow up. At a median follow up time of 9.5 years after rFNA sampling, 16 women developed invasive or non-invasive breast cancer. There were no significant differences between women who developed cancer and those that did not in regard to demographic factors, %breast density, MS, or Gail-LR. On univariate analysis, Gail-LR was higher in women who developed cancer (13.0 vs. 16.5, p=0.08). The largest hazard ratios were observed from high breast density (2.30, 95% CI 0.8, 6.6) and high CMI (2.26, 95% CI 0.8, 6.6, p=0.07). In breast cancer prediction, the AUC for CMI was 0.64 (95% CI 0.51, 0.77). In separate bivariable models that adjusted for age, Gail-LR, MS, and %breast density, the HR for log CMI was consistently above unity, with a p value consistently below 0.1, except for the model that included MS (see Table). Conclusions: Elevated CMI has potential as a robust predictor of future breast cancer occurrence in average risk women, even when adjusted for breast density or cytologic atypia. Our prior analysis established that CMI is not susceptible to variation with menstrual cycle phase and menopausal status. These features support its further evaluation in larger trials. Clinical trial information: NCT00896636 .[Table: see text]

Magnetic Resonance Imaging-Guided Breast Biopsy in Korea: A 10-Year Follow-Up Experience.

PurposeTo evaluate the accuracy of magnetic resonance imaging (MRI)-guided breast biopsy.MethodsWe retrospectively reviewed the clinical data of 111 consecutive patients referred for MRI-guided breast biopsy after mammography and breast ultrasound between May 2009 and April 2019. After excluding 37 patients without follow-up images (> 2 years), 74 patients (74 lesions) were finally included. We reviewed the histologic results of MRI-guided biopsy and subsequent surgery, post-biopsy management, and breast cancer development during follow-up. We investigated the false-negative rate, ductal carcinoma in situ (DCIS) underestimation, atypical ductal hyperplasia (ADH) underestimation rate, and technical failure rate of MRI-guided biopsy.ResultsAmong 74 scheduled MRI-guided biopsies, six were canceled because biopsy was deemed unnecessary, while three failed due to technical difficulties (technical failure rate: 3/68, 4.4%). MRI-guided biopsy was performed in 65 patients, of which 18 patients were diagnosed with malignant lesions, 46 with benign lesions, and one with ADH bordering on DCIS. Subsequent surgery (n = 27) showed DCIS underestimation in three cases (3/7, 43%), ADH underestimation in two cases (1/2, 50%), as well as seven concordant benign and 11 concordant malignant lesions. The overall false-negative rate was 4.3% (2/46). Thirty-eight out of 48 benign lesions were followed-up (median period, 5.8 years; interquartile range, 4.1 years) without subsequent surgery. Thirty-seven concordant benign lesions were stable (n = 27) or disappeared (n = 10); however, the size of one discordant benign lesion increased on follow-up MRI and it was diagnosed as DCIS after 1 year.ConclusionMRI-guided biopsy is an accurate method for exclusion of malignancy with a very low false-negative rate.

Lobular intraepithelial neoplasia: Outcomes and optimal management.

Lobular neoplasia is a term encompassing both atypical lobular hyperplasia and lobular carcinoma in situ. These pathological findings are of uncertain malignant potential and predispose to a higher lifetime risk of breast cancer. Debate surrounds the management of such lesions, with the rationale for diagnostic excision based on the possibility of upgrading to malignancy. In this study, we report the upgrade rate of these lesions and risk of subsequent development of breast cancer. This is a retrospective review of a prospectively maintained data base of all biopsies of breast screening-detected abnormalities in a single Irish breast-screening unit. We included all patients with lobular neoplasia on core needle biopsy who underwent diagnostic excision from 2005 to 2012. We excluded those who had concurrent high-risk lesions on biopsy. End points included upgrade rate and subsequent diagnosis of malignancy on follow-up. During the study period, 66 patients met criteria for inclusion, with a mean age of 53.74years. Upgrade rate following excision was 13.64% (n=9/66). Of those not upgraded, 7.02% (n=4/57) were subsequently diagnosed with malignancy. Median time to diagnosis was 59.61months (range=10.5-124.4). There is a significant rate of upgrade following diagnostic excision of lobular neoplasia, supporting the practice of diagnostic excision. There is an increased lifetime risk of breast cancer for women with a diagnosis of lobular neoplasia, with many of these cancers occurring outside the standard five-year monitoring period, suggesting a potential benefit in extending surveillance.

Abstract P1-02-06: Effect of chemoprevention on convolutional neural network-based breast cancer risk model using a mammographic dataset of women with atypical hyperplasia, lobular and ductal carcinoma in situ

Abstract Background: Mammographic density (MD) is a strong predictor of breast cancer risk and has been shown to predict response to tamoxifen for chemoprevention. We have developed a novel convolutional neural network (CNN)-derived pixel-wise breast cancer risk model, which is a more accurate predictor of breast cancer risk than MD. Our objective was to investigate how chemoprevention use among women with atypical hyperplasia (AH), lobular or ductal carcinoma in situ (LCIS/DCIS) is associated with change in the CNN breast cancer risk model. Methods: We conducted a retrospective cohort study of patients diagnosed with AH, LCIS, or DCIS at Columbia University Irving Medical Center (CUIMC) in New York, NY between 2007 and 2015. We collected mammograms conducted at baseline (at diagnosis of AH/LCIS/DCIS or start of chemoprevention) and 3-5 years later. Demographic and clinical characteristics, chemoprevention use, and mammography images were extracted from the electronic health record. Briefly, each mammogram was normalized as a map of z-scores and resized to an input image size of 256x256. Then a contracting and expanding fully convolutional CNN architecture was composed entirely of 3x3 convolutions, a total of four strided convolutions instead of pooling layers, and symmetric residual connections. L2 regularization and augmentation methods were implemented to prevent over-fitting. High-risk was defined as an absolute CNN model score &amp;gt;0.5. Change in CNN breast cancer risk model from baseline to follow-up was defined as: 1) stay high risk; 2) stay low risk; 3) increase from low to high risk; 4) decrease from high to low risk. Unordered polytomous regression models were used to assess the association between change in CNN score with chemoprevention use. Results: Among 541 evaluable patients, the mean age was 60 years (SD, 9.56) and 69% were postmenopausal. About a third of women (N=184, 34%) initiated chemoprevention, including 72% on a selective estrogen receptor modulator (SERM), 19% on an aromatase inhibitor (AI), and 9% both. About 57% had high CNN breast cancer risk scores at baseline. Comparing women who initiated chemoprevention to those who did not, more women had a decrease in their CNN breast cancer risk model (33.7% vs. 22.9%) and fewer women had an increase in CNN score (11.4% vs. 20.2%). In multivariate analysis, those who were treated with chemoprevention were less likely to have an increase in CNN risk score (odds ratio=0.51, 95% confidence interval=0.28-0.92). Conclusions: We previously reported that our CNN based algorithm can predict breast cancer risk and we now demonstrate that the CNN breast cancer risk model is modifiable on serial mammograms from high-risk women with proven chemopreventive agents, such as SERMs and AIs. Future studies should focus on whether these CNN mammographic changes are associated with subsequent development of breast cancer and may serve as a pharmacodynamics surrogate endpoint for breast cancer chemoprevention trials. Citation Format: Haley Manley, Simukayi Mutasa, Richard Ha, Katherine Crew. Effect of chemoprevention on convolutional neural network-based breast cancer risk model using a mammographic dataset of women with atypical hyperplasia, lobular and ductal carcinoma in situ [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P1-02-06.

Abstract P3-09-11: Modulation of the immune microenvironment in high risk DCIS

Abstract Background. Ductal carcinoma in situ (DCIS) is a risk factor for the subsequent development of invasive breast cancer. Features of DCIS that are associated with a high risk of a subsequent event include large size (&amp;gt; 5 cm), high grade, comedo necrosis, palpable mass, hormone receptor negativity, and HER2 positivity. We have previously shown that immune infiltrates in DCIS are positively associated with these high-risk features, suggesting that manipulating the immune microenvironment in high risk DCIS could potentially alter disease progression. Objectives. The objectives of this study were: 1). to define dose limiting toxicities, tolerability, and feasibility of intralesional injection of an immune checkpoint inhibitor (pembrolizumab) into DCIS; and 2). to determine response rate as measured by an increase in total T cells or CD8+ T cells from baseline to post treatment. Methods. Study participants received 2 intralesional injections of pembrolizumab, 3 weeks apart, with surgery approximately 3 weeks after the 2nd dose. Multiplex immunofluorescence analyses were used to compare immune cell populations in pre-treatment biopsies to post-treatment surgical specimens. Tissue samples were stained with two 6-plex immune panels using Opal immunofluorescence reagents (Perkin Elmer) on a fully automated Ventana Discovery platform, imaged with a Vectra 3 system and analyzed with inForm software (Perkin Elmer). Results. The intralesional injections were well tolerated and there were no systemic toxicities observed. Multiplex immunofluorescence analyses demonstrated significant increases in total T cells, as well as cytotoxic CD8+ T cells, following therapy. We also observed a significant positive correlation between the change (pre- vs. post-therapy) in Ki67+ T cells (proliferating T cells) with the increase in total T cells, suggesting that immune checkpoint inhibition removed the “brakes” from the T cells, allowing them to proliferate. Changes in macrophage, B cell, and Treg numbers pre- vs. post-therapy were not significant. Despite an increase in T cell infiltrates, there were no measurable indicators of an anti-tumor response: no reduction in lesion size by MRI, no reduction in proliferation of DCIS cells (Ki67 staining), and no increase in cell death (cleaved caspase 3 staining). Spatial analyses indicated that in most cases the T cell expansion (both total T cells and CD8+ T cells) occurred predominantly in the stroma, not among the DCIS cells in the ducts. Conclusions. We met the objectives of our study: 1). we demonstrated the safety and feasibility of intralesional injection of an immune checkpoint inhibitor (pembrolizumab) in high risk DCIS; and 2). we demonstrated that local immunotherapy resulted in the local proliferation/expansion of T cells. However we did not observe any change in the size of the lesions by MRI. In addition, there was no evidence for T cell killing at the cellular level, suggesting that even though there was an expansion of T cells, there are other factors at play such as immune exclusion or possibly immunosuppression of their killing ability. These results suggest that combination of pembrolizumab with another agent(s) will be needed to achieve a more complete response. Citation Format: Michael Campbell, Jasmine Wong, Emma McCune, Harriet Rothschild, Tristan Loveday, Jennifer Bolen, Cliff Hoyt, Scott VandenBerg, Jo Chien, Laura Esserman. Modulation of the immune microenvironment in high risk DCIS [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P3-09-11.

Flat epithelial atypia on core needle biopsy does not always mandate excisional biopsy

Flat Epithelia Atypia (FEA) is a proliferative lesion of the breast where cells demonstrate columnar change and cytologic atypia. This lesion has been identified as distinct from the classic atypical hyperplasias (AH). While many patients undergo excisional biopsy, management of FEA identified on core needle biopsy (CNB) is controversial, and the rate of associated ductal carcinoma in situ (DCIS) or invasive cancer is not well defined. The aim of this study was to determine the upstage rate of FEA diagnosed by CNB. We identified patients from a prospectively maintained data base who had FEA diagnosed by CNB from 01/2010 to 07/2015. Patient variables collected included age at presentation, imaging findings, pathologic findings following surgical excision, and subsequent development of breast cancer. Of 132 patients, 62 (n=62/132, 47.0%) patients had FEA associated with DCIS and invasive ductal carcinoma (IDC) on CNB and were excluded from analysis. Of the remaining 70 patients, median age was 52 (range 31-84) years. Thirty-two (45.7%) patients had FEA plus AH, 4 (5.7%) patients had FEA plus lobular carcinoma in situ (LCIS), and 34 (48.6%) patients had FEA alone or with another non-pathologic finding (pure FEA). Two (6.3%) patients with FEA plus AH had DCIS or IDC on subsequent excisional biopsy. Of the 34 patients with pure FEA who underwent excisional biopsy, only one (2.9%) was found to have IDC. Twenty-two (64.7%) patients with pure FEA who underwent excisional biopsy presented with calcifications on mammography. None of these patients had cancer on excisional biopsy, and 10 (45.5%) patients had AH (3 ADH, 3 ALH, and 4 both ALH and ADH). Twelve (n=12/34, 35.3%) patients with pure FEA underwent CNB for a mass or asymmetry noted on imaging. Of these 12 patients, 9 (75.0%) had benign findings on excisional biopsy, two (16.7%) patients had AH, and one (8.3%) patient had IDC. Median follow-up was 4.6years (IQR 3.1-6.5years). Three (4.3%) patients subsequently developed IDC, two of which were in the contralateral breast. FEA is often found in combination with ADH and ALH as well as carcinoma on CNB. In our study, pure FEA was upstaged to cancer in only 2.9% of patients. Mammographic findings unlikely predict upstaging to malignancy. These findings suggest that excisional biopsy may not be warranted in patients with pure FEA and could be managed with close imaging surveillance.

Abstract P2-09-02: Modulation of the immune microenvironment in high risk DCIS by intralesional injection of anti-PD-1 (pembrolizumab)

Abstract Background. Ductal carcinoma in situ (DCIS) is a risk factor for the subsequent development of invasive breast cancer. Features of DCIS that are associated with a high risk of a subsequent event include large size (&amp;gt; 5 cm), high grade, comedo necrosis, palpable mass, hormone receptor negativity, and HER2 positivity. We have previously shown that immune infiltrates are positively associated with these high-risk features, suggesting that manipulating the immune microenvironment in high-risk DCIS, for example via checkpoint blockade, could potentially alter disease progression. Methods. In this phase 1 pilot study we investigated changes in the immune microenvironment of high risk DCIS after intralesional injection of anti-PD-1 (pembrolizumab). Study participants received 2 intralesional injections of pembrolizumab, 3 weeks apart, with surgery approximately 3 weeks after the 2nd dose. The study started with a dose of 2 mg/injection (1/100 of the standard 200 mg iv dose), then escalated to 4 mg and 8 mg, with 3 patients at each dose. Tissue samples from pre-treatment biopsies and post-treatment surgical resections were stained with two 6-plex immune panels using Opal immunofluorescence reagents (Perkin Elmer) on a fully automated Ventana Discovery platform, imaged with a Vectra 3 system and analyzed with inForm software (Perkin Elmer). An algorithm for tumor/stroma segmentation developed in inForm was used to randomly select 10 high power fields (hpfs) for imaging. Cell phenotype maps were generated for each of these hpfs for each sample. Cell densities were determined per area of stroma, DCIS, or total tissue and averaged across all hpfs for a given case. Spatial analyses were performed to quantitate co-localization of immune cells with DCIS cells. Results. The intralesional injections were easily administered and well tolerated. No systemic toxicities were observed at any dose. MRI imaging demonstrated no change in the size of lesions after treatment. Multiplex immunofluorescence analyses demonstrated heterogeneous responses ranging from dramatic increases in T cells, in particular CD8+ T cells, in cases which had a T cell infiltrate prior to therapy, to no post-therapy T cell infiltrate in cases with a pre-therapy immune desert. We also observed increases in B cells and macrophages and a decrease in the ratio of FoxP3+ T cells to CD8+ T cells, the latter mainly due to a significant increase in CD8+ cells, as opposed to a decrease in FoxP3+ cells. Spatial analyses indicated that in some cases, despite a marked increase in T cells post therapy, these cells did not co-localize with DCIS cells, indicating a state of immune exclusion. Conclusions. We have demonstrated the safety and feasibility of intralesional injection of an immune checkpoint inhibitor (pembrolizumab) in high risk DCIS. In some patients we observed a dramatic change in the immune microenvironment, with an increase in T cells, B cells, and macrophages, and a decrease in the FoxP3:CD8 ratio, even at a dose that is 1/100 of the standard intravenous clinical dose. An expansion study is underway in which patients will receive 4 injections of pembrolizumab at 3 week intervals prior to going to surgery to determine if more injections/time will increase response rate. Citation Format: Campbell MJ, McCune E, Rothschild H, Bolen J, VandenBerg S, Chien J, Wong J, Esserman L. Modulation of the immune microenvironment in high risk DCIS by intralesional injection of anti-PD-1 (pembrolizumab) [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 P2-09-02.

Negative Association of Proton Pump Inhibitors With Subsequent Development of Breast Cancer: A Nationwide Population-Based Study.

Although current evidence suggests potential antitumor activity of proton pump inhibitors (PPIs), there is no population-based evidence of an association between PPI use and subsequent breast cancer risks. We used an observational case-control study to examine the association between prior PPI use and breast cancer occurrence. Additional analysis examined dose-response and age-stratified associations of PPIs with breast cancer. This study used data from the Taiwan National Health Insurance Research Dataset. A total of 64,234 women diagnosed with breast cancer between 2004 and in 2013 were selected as cases. Controls were 64,234 women without cancer who were selected by matching them with cases on the basis of sociodemographic characteristics and widely prevalent comorbidities. Each study subject's claims data were tracked back for 5 years to determine precancer prescriptions of PPIs. Logistic regression modeling was used for the analysis. A total of 11,871 (9.24%) women had used PPIs within the prior 5 years, 8.06% and 10.42% among cases and controls, respectively. Breast cancer patients were 25% less likely to have had prior PPI exposure after adjustment for comorbidities that predispose to PPI exposure (95%CI 0.72-0.78) in the risk of breast cancer occurrence. A dose-response effect was also detected, with the highest effect, 35% lower PPI odds (95%CI 0.61-0.70) among patients in the highest exposure category. Our findings may suggest that women at a higher-than-average risk of breast cancer may benefit from PPI prescriptions if they have medical conditions that could benefit from PPIs.

Breast cancer risk associated with atypical hyperplasia and lobular carcinoma in situ initially diagnosed on core-needle biopsy.

Breast cancer risk estimates for atypical lesions are based primarily on case-control studies of patients with open biopsies. The authors report the cumulative breast cancer incidence after a core biopsy diagnosis of atypical hyperplasia (ductal or lobular) or lobular carcinoma in situ. A cohort study with central pathology review was conducted on 393 patients who had core biopsy diagnoses of atypical hyperplasia and lobular carcinoma in situ from 1995 through 2010. Follow-up was available for 255 of 264 patients (97%) at a median of 87 months (range, 3-236 months). There were 212 patients (54%) who were not upgraded on excision and had no personal history of breast cancer. Of these, 21 of 212 (9.9%) developed breast cancer, including 15 invasive carcinomas, 4 ductal carcinomas in situ, 1 pleomorphic lobular carcinoma in situ, and 1 unknown type. The prior core biopsy diagnoses were atypical ductal hyperplasia for 11 patients (52%) and atypical lobular hyperplasia/lobular carcinoma in situ in the remaining 10 patients (48%). The number of atypical foci in the core biopsy was not significantly associated with the subsequent development of breast cancer (P = .42). Of the 15 invasive carcinomas, 11 (73%) were ipsilateral, 11 (73%) were pathologic T1 tumors, 5 (33%) were pathologic N1 tumors, 13 (87%) were estrogen receptor-positive, and 1 (7%) was amplified for human epidermal growth factor receptor 2. In patients who had an initial diagnosis of atypical hyperplasia or lobular carcinoma in situ on core biopsy, the 7-year cumulative breast cancer incidence was 9.9%. Most tumors were ipsilateral, stage I, estrogen receptor-positive, invasive carcinomas. The current data support close clinical and radiologic follow-up for more than 5 years in this patient population. Cancer 2018;124:459-65. © 2017 American Cancer Society.

Aromatase expression in atypical ductal hyperplasia in women.

To determine the levels of aromatase in atypical ductal hyperplasia (ADH) lesions, tissue surrounding the ADH, and in dense and non-dense normal breast tissue. We postulated that excess aromatase in breast tissue might, through production of increased estrogen, drive the carcinogenic process. Estrogens and their metabolites are thought to contribute to the development of breast cancer through estrogen receptor-mediated mechanisms and genotoxic effects of estrogen metabolites. ADH is a benign lesion of the breast which is associated with substantially increased risk for subsequent development of breast cancer. After 25 years, approximately 30% of women with ADH develop breast cancer. In women with three or more separate ADH lesions at the same time, 47% will develop breast cancer over that time period. Another important risk factor for breast cancer is the presence of mammographically dense breast tissue. We utilized quantitative immunochemical analysis of aromatase in biopsy tissue to test this possibility. Previously published results comparing dense with non-dense breast tissue in normal women (Vachon et al. Breast Cancer Res Treat 125:243-252, 2011) were used for comparisons with ADH. A well-characterized histochemical H-score was employed for quantitative assessment of aromatase in the various tissue studied. The H-score of aromatase staining was statistically significantly higher (p = 0.003) in the ADH epithelium than surrounding epithelial tissue. In order of H-score from highest to lowest were ADH, issue surrounding ADH, dense normal and non-dense normal breast tissues. The levels of aromatase in a subset of women with ADH who went on to develop breast cancer were not higher than in women who did not. We suggest from these studies that overexpression of aromatase in breast tissue and its resultant increase in estradiol levels may contribute to the later development of breast cancer in women with ADH.

Abstract LB-364: Natural history of age-related lobular involution and impact on breast cancer risk

Abstract Background: Age-related lobular involution (LI) is a physiological process in which the terminal duct lobular units of the breast regress as a woman ages. Analyses of breast biopsies from women with benign breast disease (BBD) have found that extent of LI is negatively associated with subsequent breast cancer development. Here we assess the natural course of LI within individual women, and the impact of progressive LI on breast cancer risk. Methods: The Mayo Clinic BBD cohort consists of 13,455 women with BBD from 1967-2001. The BBD cohort includes 1115 women who had multiple benign biopsies, 106 of whom had developed breast cancer. Within this multiple biopsy cohort, the progression of the LI process was examined by age at initial biopsy and time between biopsies. The relationship between LI progression and breast cancer risk was assessed using standardized incidence ratios (SIRs) and by Cox proportional hazards analysis. Results: Women who had multiple biopsies were younger age and had a slightly higher family history of breast cancer as compared with the overall BBD cohort. Extent of LI at subsequent biopsy was greater with increasing time between biopsies and for women age 55+ at initial biopsy. Among women with multiple biopsies, there was a significant association of higher breast cancer risk among those with involution stasis (lack of progression, HR 1.63) as compared with those with involution progression, p = 0.036. Conclusions: The multiple biopsy BBD cohort allows for a longitudinal study of the natural progression of LI. The majority of women in the multiple biopsy cohort showed progression of LI status between benign biopsies, and extent of progression was highest for women who were in the perimenopausal age range at initial biopsy. Progression of LI status between initial and subsequent biopsy was associated with decreased breast cancer risk. Citation Format: Derek Radisky, Daniel W. Visscher, Ryan Frank, Robert A. Vierkant, Stacey J. Winham, Melody L. Stallings-Mann, Tanya L. Hoskin, Aziza Nassar, Celine M. Vachon, Lori A. Denison, Lynn C. Hartmann, Marlene H. Frost, Amy C. Degnim. Natural history of age-related lobular involution and impact on breast cancer risk. [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 LB-364.

Abstract LB-378: Ankyrin repeat domain 30A as a novel molecular marker of columnar alterations in benign breast disease and subsequent breast cancer in African American women

Abstract An estimated 1.6 million breast biopsies are performed annually in the United States, and the vast majority are benign. Benign breast disease is an established breast cancer risk factor for both Caucasian and African American women. Yet little is still known about benign breast tissue from African American women, who suffer from disproportionately high-grade tumors and poorer breast cancer survival. To better characterize the risk of breast cancer among African American women with benign breast disease, we assessed benign breast disease pathology of African American women in our metropolitan Detroit cohort (n = 3,737). We classified benign breast lesions for pathological characteristics and used logistic regression to estimate subsequent breast cancer risk with histological characteristics. We observed that women whose biopsies showed columnar alterations, columnar cell lesions of the breast, were 1.78-fold more likely to develop an invasive breast cancer [odds ratio (OR) 1.78, 95% confidence interval (CI) 1.19-2.65] after adjusting for age at biopsy, biopsy year, and overall impression. Columnar alterations were not found to be significantly associated with risk of developing ductal carcinoma in situ (DCIS) among African American women [OR 1.53, 95% CI 0.83-2.82]. Next, to identify molecular markers associated with columnar alterations in benign breast lesions, we performed gene expression analyses on biopsy tissues from 27 African American women that subsequently developed an invasive breast cancer. Transcriptional profiling of these lesions identified differentially expressed genes significantly associated with columnar alterations, including ankyrin repeat domain 30A (ANKRD30A). ANKRD30A, also known as NY-BR-1, is a mammary tissue-specific differentiation antigen that is overexpressed in breast cancers. Increased expression of ANKRD30A in benign breast lesions with columnar alterations may be indicative of subsequent breast cancer development. Taken together, novel molecular markers, including ANKRD30A, and ethnicity can lead to better risk assessment and earlier diagnosis of breast cancer. Citation Format: Andreana N. Holowatyj, Rouba Ali-Fehmi, Julie J. Ruterbusch, Eman Abdulfatah, Vishakha Pardeshi, Woodlyne Roquiz, Baraa Alosh, Sudeshna Bandyopadhyay, Derek Radisky, Michele L. Cote. Ankyrin repeat domain 30A as a novel molecular marker of columnar alterations in benign breast disease and subsequent breast cancer in African American women. [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 LB-378.

Natural history of age-related lobular involution and impact on breast cancer risk.

Age-related lobular involution (LI) is a physiological process in which the terminal duct lobular units of the breast regress as a woman ages. Analyses of breast biopsies from women with benign breast disease (BBD) have found that extent of LI is negatively associated with subsequent breast cancer development. Here we assess the natural course of LI within individual women, and the impact of progressive LI on breast cancer risk. The Mayo Clinic BBD cohort consists of 13,455 women with BBD from 1967 to 2001. The BBD cohort includes 1115 women who had multiple benign biopsies, 106 of whom had developed breast cancer. Within this multiple biopsy cohort, the progression of the LI process was examined by age at initial biopsy and time between biopsies. The relationship between LI progression and breast cancer risk was assessed using standardized incidence ratios and by Cox proportional hazards analysis. Women who had multiple biopsies were younger age and had a slightly higher family history of breast cancer as compared with the overall BBD cohort. Extent of LI at subsequent biopsy was greater with increasing time between biopsies and for women age 55 + at initial biopsy. Among women with multiple biopsies, there was a significant association of higher breast cancer risk among those with involution stasis (lack of progression, HR 1.63) as compared with those with involution progression, p = 0.036. The multiple biopsy BBD cohort allows for a longitudinal study of the natural progression of LI. The majority of women in the multiple biopsy cohort showed progression of LI status between benign biopsies, and extent of progression was highest for women who were in the perimenopausal age range at initial biopsy. Progression of LI status between initial and subsequent biopsy was associated with decreased breast cancer risk.Electronic supplementary materialThe online version of this article (doi:10.1007/s10549-016-3691-5) contains supplementary material, which is available to authorized users.

Gene signature model for breast cancer risk prediction for women with sclerosing adenosis.

18 Background: Benign breast disease (BBD) is diagnosed in 1-2 million women/year in the US, and while these patients are known to be at substantially increased risk for subsequent development of breast cancer, existing models for risk assessment perform poorly at the individual level. Here, we describe a DNA-microarray-based transcriptional model for breast cancer risk prediction for patients with sclerosing adenosis (SA), a lesion found in ¼ of all BBD patients that is characterized by epithelial proliferation, disordered acinar architecture, and stromal fibrosis. Methods: A training set was developed from 86 patients diagnosed with sclerosing adenosis (SA), of which 27 subsequently developed cancer within 10 years (cases) and 59 remained cancer-free at 10 years (controls). A diagonal linear discriminate analysis (DLDA)-prediction model for prediction of cancer within 10 years (SA TTC10) was generated from transcriptional profiles of formalin fixed, paraffin-embedded (FFPE) biopsy-derived RNA. This model was tested on a separate validation case-control set composed of 65 SA patients. Results: The SA TTC10 gene signature model, composed of 35 gene features, achieved a clear and significant separation between case and control with receiver operating characteristic area under the curve of 0.913 in the training set and 0.836 in the validation set. Conclusions: Our results provide the first demonstration that benign breast tissue contains transcriptional alterations that indicate risk of breast cancer development, and that essential precursor biomarkers of malignancy are present many years prior to cancer development. Furthermore, the SA TTC10 gene signature model, which can be assessed on FFPE biopsies, constitutes a novel prognostic biomarker for patients with SA.