Visscher, Daniel W.

1061 Background: Benign breast disease (BBD) is a significant risk factor for breast cancer (BC); however little is known about the aggressiveness of the BCs these patients develop. Shared features between select atypias and low-grade DCIS have led some to speculate that a history of BBD portends the development of well-differentiated BCs. Methods: The Mayo BBD cohort includes 9,376 women who had benign breast biopsies from 1967–1991. Data on BC events were obtained from medical records and questionnaires. Tissue from the BCs was reviewed by a single breast pathologist (DWV). Results: Our cohort included 6,244 patients with nonproliferative disease (NP), 2,801 women with proliferative disease without atypia (PDWA), and 331 with atypical hyperplasias (AH). With a median of 18 years of follow-up, 799 patients with BBD have developed breast cancer, 416 initially had NP, 313 had PDWA, and 70 AH. BC tissue was available for 703 of the women. The cancers were invasive ductal in 76.1% (n = 535), invasive lobular in 9.4% (n = 66), and DCIS alone in 14.5% (n = 102) patients. Grade is currently available for 500 women, and is well differentiated in 29.4% (n = 150), moderately differentiated in 42.7% (n = 218) and poorly differentiated in 27.8% (n = 142). In 537 cases the malignant tissue had a concurrent benign component consisting of NP in 24% (n = 129), PDWA in 25.1% (n = 135), and AH in 50.8% (n = 273). In regards to tumor size, 69.6% had T1 tumors, 24.9% T2 tumors, and 5.5% T3 disease. Metastasis to lymph nodes occurred in 25% of patients. Median time to BC was 12.3 years and did not differ across the different benign entities. Breast cancer developed within 5 years from the initial BBD in 163 women (20%). In this group, tumor size was greater (p = 0.02), and there were more poorly differentiated tumors (p = 0.006) than in those who later developed BC. Conclusions: In the BCs that have developed in our BBD cohort, we do not see a preponderance of well-differentiated, good-risk BCs. Patients diagnosed within 5 years of BBD appear to have BCs with more aggressive features. No significant financial relationships to disclose.

Abstract Over one million women in the United States receive biopsy diagnoses of benign breast disease (BBD) each year, which confer a 1.5–4.0-fold increase in breast cancer risk. Studies in the general population suggest that nonsteroidal anti-inflammatory agents (NSAID) lower breast cancer risk; however, associations among women with BBD are unknown. We assessed whether NSAID use among women diagnosed with BBD is associated with lower breast cancer risk. Participants included 3,080 women (mean age = 50.3 ± 13.5 years) in the Mayo BBD surgical biopsy cohort diagnosed between January 1, 1992 and December 31, 2001 who completed breast cancer risk factor questionnaires that assessed NSAID use, and whose biopsies underwent detailed pathology review, masked to outcome. Women were followed from date of BBD biopsy to breast cancer diagnosis (main outcome) or censoring (death, prophylactic mastectomy, reduction mammoplasty, lobular carcinoma in situ or last contact). Median follow-up time was 16.4 ± 6.0 years. Incident breast cancer was diagnosed among 312 women over a median follow-up of 9.9 years. Regular non-aspirin NSAID use was associated with lower breast cancer risk [HR = 0.63; 95% confidence interval (CI) = 0.46–0.85; P = 0.002] with trends of lower risk (highest tertiles of use vs. nonuse) for greater number of years used [HR = 0.55; 95% CI = 0.31–0.97; Ptrend = 0.003), days used per month (HR = 0.51; 95% CI = 0.33–0.80; Ptrend = 0.001) and lifetime number of doses taken (HR = 0.53; 95% CI = 0.31–0.89; Ptrend = 0.003). We conclude that nonaspirin NSAID use is associated with statistically significant lower breast cancer risk after a BBD biopsy, including a dose–response effect, suggesting a potential role for NSAIDs in breast cancer prevention among patients with BBD.

Abstract Treatment of ovarian cancer is limited by extensive metastasis and yet it remains poorly understood. We have studied the critical step of metastatic colonization in the context of the productive interactions with the metastatic microenvironment with a goal of identifying key regulators. By combining miRNA expression analysis using an organotypic 3D culture model of early ovarian cancer metastasis with that of matched primary and metastatic tumors from 42 patients with ovarian cancer, we identified miR-4454 as a key regulator of both early colonization and advanced metastasis in patients with ovarian cancer. miR-4454 was downregulated in the metastasizing ovarian cancer cells through paracrine signals from microenvironmental fibroblasts, which promoted migration, invasion, proliferation, and clonogenic growth in ovarian cancer cells as well as their ability to penetrate through the outer layers of the omentum. Stable overexpression of miR-4454 decreased metastasis in ovarian cancer xenografts. Its mechanism of action was through the upregulation of its targets, secreted protein acidic and cysteine rich (SPARC) and BCL2 associated athanogene 5 (BAG5), which activated focal adhesion kinase (FAK) signaling, promoted mutant p53 gain of function by its stabilization, and inhibited apoptosis. Because microenvironment-induced downregulation of miR-4454 is essential for early and advanced metastasis, targeting it could be a promising therapeutic approach. Implications: This study identifies a miRNA, miR-4454, which is downregulated by signals from the microenvironment and promotes early and advanced ovarian cancer metastasis through its effects on FAK activation, mutant p53 stabilization, and apoptosis inhibition.

Abstract Background: TNBC is the most aggressive sub-type of breast cancer and has limited targeted therapies. Atezolizumab was recently FDA-approved for patients with advanced PD-L1+ TNBC. In a well-characterized cohort of patients with early-stage TNBC, we used the FDA-approved PD-L1 SP142 companion assay for atezolizumab to score whole tumor sections for PD-L1 expression. We determined the frequency and scoring distribution of PD-L1 in these tumors and evaluated PD-L1 status for associations with clinicopathologic variables and outcomes. Methods: Whole tumor sections from 498 women with surgically-treated, non-(distant) metastatic TNBC (ER/PR <1% & HER2-negative by central review) were semi-quantitatively scored for % of PD-L1-staining tumor cells (TC) or tumor-associated immune cells (IC) (PD-L1 SP142, Ventana, performed in CAP/CLIA-certified clinical laboratory). Per FDA assay guidelines, PD-L1+ tumors were defined as ≥1% of tumor-associated immune cells (IC) staining per tumor area. Baseline clinical characteristics, histopathologic features, Ki-67 proliferative indices (PI), and stromal tumor infiltrating lymphocytes (TILs) were correlated with PD-L1 status. The impact of tumoral PD-L1 status on invasive disease-free survival (IDFS) and overall survival (OS) was assessed using Cox proportional hazards models. Results: The majority of tumors were pT1-T2 (95%), pN0-1 (85%), and grade 3 (90%) with Ki-67 >15% (80%). 228/498 (46%) tumors were PD-L1+, distributed as IC score 1-5%: 21%, 6-10%: 13%, 11-20%: 5% and >20%: 7%. 270/498 (54%) were PD-L1-. 43 (8.6%) of tumors had PD-L1 staining in TC (all but 1 were also IC+). PD-L1 IC expression was associated with larger tumor size (p = 0.005), higher nodal stage (≥pN1: 43% vs. 31%, p < 0.001), higher grade (grade 3: 95% vs. 86%, p< 0.001), higher Ki-67 PI (>15%: 88% vs. 69%, p <0.001), and stromal TILs (median score: 40% vs. 10%; p < 0.001). PD-L1 IC status was not associated with patient age. Among histologic subtypes, PD-L1+ tumors were more frequently medullary type (31% vs. 6%) and less frequently metaplastic type (4% vs 10%). At time of this analysis, median follow-up for IDFS and OS were 11 years and 7.2 years, respectively. By univariable analysis, absence of PD-L1 IC expression was associated with poorer IDFS (median: 15.5 vs. 20 years, p = 0.04) and OS (9.5 vs. 17.5 years, p < 0.05).By multivariable analysis, tumor size, stromal TILs, nodal status, and adjuvant chemotherapy (CTX) were significantly associated with IDFS. Age, stromal TILs, nodal status, and receipt of adjuvant CTX were associated with OS. PD-L1+ tumors trended toward improved IDFS (statistical significance not reached), HR: 0.68 (95% CI: 0.45-1.03, p= 0.068) but not OS: 0.90 (95% CI: 0.52-1.57, p= 0.32). Conclusion: Using the PD-L1 SP142 companion assay, almost half of early-stage TNBC were PD-L1+ (IC). However, PD-L1 expression correlated with increasing tumor size and nodal burden, as well as histologic grade, Ki-67 PI, stromal TILs and lymphocyte-enriched subtypes. PD-L1 was associated with improved IDFS and OS in univariable analysis but not multivariable analysis. The majority of the PD-L1+ tumors (73%) had IC scores ≤10%, with heterogeneous tumoral staining. Given the low (1%) assay threshold for positivity, these data emphasize the utility of evaluating PD-L1 in whole tumor sections rather than tissue cores. Citation Format: Jodi M Carter, Mei-Yin C Polley, Jason P Sinnwell, Roberto A Leon-Ferre, Fergus J Couch, Krishna R Kalari, Judy C Boughey, Minetta C Liu, Daniel W Visscher, Matthew P Goetz. Frequency, characteristics and prognostic factors of PD-L1+ triple negative breast cancer using the PD-L1 SP142 companion assay [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 PD1-08.

Abstract Background Prior epidemiologic studies have shown an inverse relationship of aspirin and NSAID use with breast cancer (BC) risk in the general population. Women with benign breast disease (BBD) are at an increased BC risk compared with women in the general population, and we have shown that BBD biopsies contain increased numbers of immune cells compared with normal breast tissues donated for research in the Komen Tissue Bank. We hypothesized that NSAID use among women with BBD is associated with reduced risk of BC. Methods We evaluated BC risk in relation to aspirin and NSAID use in a single institution retrospective cohort of 4,498 women who received a biopsy diagnosis of BBD between 1/1/1992 and 12/31/2001. Information on usual medication use, including average number of days used per month (0, 1-7, 8-28 and >28), was collected via questionnaires mailed after identification of participants. NSAIDs were categorized into three classes: 1) Aspirin, 2) Ibuprofen (plus other prostaglandin synthesis inhibitors), and 3) COX inhibitors. BC events were captured via medical record, questionnaires, and Tumor Registry records. Medication use was evaluated for associations with clinical and pathology features. Hazard ratios (HR) and 95% confidence intervals (CI) examining associations between medication use and BC risk were estimated using Cox regression analyses, with adjustment for age and BMI at biopsy, years between biopsy and questionnaire, and pathologic features related to BC risk (severity of BBD and degree of involution in background lobules). Tests for trend examining frequency of use were calculated from ordered values (lowest to highest) in Cox models with a one degree-of-freedom linear term. We assessed BC associations with any type of NSAID use, as well as the three subclasses. Results Information on NSAID use was available for 3,089 (69%) of the participants in the cohort; these women had a median age of 49 and did not differ in BC risk from women who did not provide data on NSAID use (p=0.46). Of the 3,089 respondents, 313 (10.1%) subsequently developed BC with median follow-up of 17 years. Age was significantly associated with use of all types of NSAIDs, with increasing use in older women. BMI at the time of biopsy, severity of BBD findings, and involution in background lobules were also associated with NSAID use; thus these factors and age were adjusted for in risk association analyses. Ever users of NSAIDs (any type) had a significantly lower risk of BC than never users (HR 0.76, CI 0.59-0.98, p=0.03), and increasing frequency of NSAID use was significantly associated with lower BC risk (HRs 0.81, 0.79, and 0.70 for regular use 1-7, 8-28, and >28 days/month), trend p=0.02. Analyses of drug classes revealed that ever use of ibuprofen (HR 0.61, CI 0.42-0.89, p=0.01) and ever use of COX inhibitors (HR 0.59, CI 0.39-0.89, p=0.01) were associated with decreased BC risk. COX inhibitors in particular showed a strong dose response effect with increasing use (HRs 0.97, 0.52, and 0.47 for regular use 1-7, 8-28, and >28 days/month), trend p<0.01. Use of aspirin was not associated with BC risk (HR 0.88, CI 0.64-1.21, p=0.44). Conclusions In this exploratory analysis, we found use of ibuprofen and other NSAIDs to be associated with decreased BC risk among women who had undergone a BBD biopsy. These findings suggest that anti-inflammatory approaches may have value in chemoprevention among women with BBD. Citation Format: Amy Degnim, Robert Vierkant, Stacey Winham, Marlene Frost, Daniel Visscher, Jodi Carter, Suneetha Kaggal, Julie Cunningham, Teresa Allers, Tanya Hoskin, Ethan Heinzen, Celine Vachon, Keith Knutson, Keith Knutson, Derek Radisky, Mark Sherman. NSAID use and breast cancer risk in a retrospective cohort of women with benign breast disease [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 P5-08-06.

Abstract Background The Circulating Cell-free Genome Atlas (CCGA; NCT02889978) study is a prospective, multi-center, observational, case-control study with longitudinal follow-up to support the development of a plasma cfDNA-based multi-cancer early detection assay. We previously reported that the fraction of tumor-derived cfDNA fragments in plasma (estimated tumor fraction [TF]) is associated with detection of multiple cancer types using a prototype whole-genome bisulfite sequencing (WGBS) assay in a pre-specified substudy. In addition, detectability of breast cancer varied by hormone-receptor (HR) status: HR negative (HR-) breast cancer had a higher detection rate than the overall breast cancer cohort. Here, we performed a planned in-depth analysis of the breast cancer cohort to understand the main clinical and biological determinants of detectability using our WGBS assay. Methods Blood samples were prospectively collected from participants (pts) with newly-diagnosed, untreated breast cancer. In a pre-specified CCGA substudy, 511 (31.4%) of 1,628 pts had a clinicopathologic diagnosis of breast cancer (any stage, excluding stage 0) and a WGBS result. Pre-specified biological and clinical factors were assessed for correlation with TF: clinical stage, pathologic N stage, lesion size (by imaging), T stage, histologic grade, hormone receptor (HR) status, and proliferative rate (Ki-67 by IHC [MIB-1 clone]) via univariate and multivariate analyses. Ki-67 was obtained from pathology reports provided by enrollment sites and also assessed centrally (Mayo Clinic [Rochester, MN]). Stage-specific differences in detection rates between HR positive (HR+) and HR- breast cancers due to biological differences such as histologic grade and/or proliferative rate was also assessed. Results Of 511 pts, 94 (18.4%; 48.9% HR- and 12.3% in HR+) had WGBS-detected cancer. Higher TF was significantly associated with cancer detection (p<0.001). In univariate analyses, the variables significantly associated with TF were clinical stage, pathological N stage, and histologic grade (p<0.001 each). Primary lesion size was modestly correlated with TF (all stages combined: Spearman's correlation coefficient rs=0.4, p<0.001), and when grouped by early and late stages (stages I/II: rs=0.27, p<0.01; stages III/IV: rs=0.50, p<0.01); clinical T stage was significantly associated with TF only when all stages were combined (p<0.001). Proliferative rate (Ki-67) was positively correlated with TF at stages I/II (rs=0.29, p<0.05) but not III/IV (rs=-0.24, p=0.34); Ki-67 results obtained through Mayo were similar to that from enrollment sites (Pearson’s correlation coefficient rp=0.68, p<0.001). HR- breast cancers had significantly higher TF versus HR+ breast cancers only at stage II (p<0.001). In addition, compared to HR+ breast cancers, HR- breast cancers had a significantly higher proportion of high histologic grade (p<0.001 at stages I and II) and proliferative rate at earlier stages (p<0.05 at stages I, II, and III). In the multivariate analysis, clinical stage (stage III vs I and IV vs I) and HR status remained the most significantly associated with TF (p<0.001 each). Conclusions Using a prototype methylation-based plasma cfDNA cancer detection assay, TF was found to be associated with detection, and with clinical and biological features of breast cancer. Features routinely used to assess clinical aggressiveness (eg, HR- status and histologic grade) also demonstrated stage-specific associations with TF. Given the known relationship between TF and detection, these data suggest that clinical and biological cancer features may also provide insight into the variability of cancer detection using plasma-based cfDNA tests. Citation Format: Minetta C. Liu, Jodi M. Carter, Daniel W. Visscher, Karla Kopp, Rita Shaknovich, Xiaoji Chen, Kathryn N. Kurtzman, Shilpen Patel, Jacqueline D. Brooks, Carlo R. Cosenza, Jafi A. Lipson, Donald A. Richards, Fergus J. Couch, Zhao Dong, Hai Liu, Oliver Venn, Joerg Bredno, Eric T. Fung, Anne-Renee Hartman. Blood-based cancer detection in plasma cell-free DNA (cfDNA): Evaluating clinical and pathologic tumor characteristics in participants with breast cancer [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 P5-01-01.

5546 Background: CHFR, an E3 ubiquitin ligase that regulates Aurora A and Pololike Kinase 1, plays a critical role in the cellular response to mitotic stress. In particular, cells containing CHFR arrest in G2 when microtubule dynamics are disrupted. Conversely, low CHFR expression is associated with hypersensitivity to mitotic spindle poisons in breast cancer cell lines. Despite the extensive use of taxanes in front line treatment of epithelial ovarian cancer (EOC), little is known about CHFR expression in this disease. We examined CHFR expression and its relationship with clinical characteristics and outcome in women with EOC. Methods: Pretreatment EOC samples from women enrolled in the Mayo Clinic Biospecimen Resource for Ovarian Cancer Research who underwent initial surgical debulking between 1999-2009 were stained in triplicate with anti-CHFR antibodies. Samples were scored in a blinded fashion for CHFR expression. Associations between CHFR staining and histology, grade, and stage were assessed using chi-squared tests; associations between CHFR and overall survival (OS) or time to disease recurrence (TTR) were assessed using Cox proportional hazards models. Results: Samples from 354 women were stained; median age at diagnosis was 60.5 (range 21-93). At a median follow-up of 67 months, 221 women (62%) had died. Across all patients, moderate/strong CHFR staining was associated with poor OS (HR = 1.39, 95% CI: 1.07-1.81, p = 0.015). Moreover, moderate/strong CHFR expression was strongly associated with serous histology (p < 0.0001), high grade (p < 0.0001) and advanced stage (p = 0.0002). To assess whether the association between CHFR expression and OS was related to taxane sensitivity, a subgroup analysis was performed in the subset of patients (N=131) with high-grade serous EOC who received initial platinum/taxane chemotherapy. Moderate/strong CHFR expression failed to correlate with OS (HR=0.91, 95% CI: 0.57-1.45, p = 0.68) or TTR from the start of chemotherapy (HR = 0.85, 95% CI: 0.55-1.31, p = 0.55) in this subset. Conclusions: CHFR expression is associated with poor OS and adverse clinical characteristics in patients with EOC but does not appear to be related to taxane sensitivity in high-grade serous tumors.

2562 Background: Empiric chemotherapy (taxane/platinum) is standard for CUP. Because prognosis is poor, novel approaches are needed. The PI3K/mTOR pathway is frequently dysregulated in cancer. Everolimus (E), an mTOR inhibitor, is approved for multiple malignancies. We performed a phase II study of Pac + Carb + E as first-line therapy in metastatic CUP patients (pts). We additionally determined if a gene expression profiling (GEP) test that identifies tissue of origin (Pathwork Tissue of Origin) could identify responsive pts. (NCT00936702) Methods: Newly diagnosed, untreated CUP pts were eligible. Central pathology review confirmed CUP prior to registration; GEP was performed on formalin fixed tumor tissue. Pac (200 mg/m2), Carb (AUC=6) and E (30 mg once weekly) were delivered every 3 wks until progression or intolerable adverse events (AEs). The primary endpoint was confirmed response, with ≥11 of 50 responses (22%) needed for trial success. Secondary endpoints were OS, progression-free survival (PFS) and AEs. Results: 46 pts (median age 61) received a median 4 cycles (range: 1-33). 39 (85%), 21 (46%) and 1 (2%) experienced ≥1 grade (gr) 3+, 4+, or 5 (sepsis) AE, with gr 3+ hematologic AE most common (74%). Of 44 evaluable pts, 15 had a confirmed response (RR 34%, 95% CI: 21-50%), with a median PFS and OS of 4.1 and 10.1 mos, respectively. Adequate tissue for GEP was available in 36 pts and predicted 10 different sites of origin. In pts with a predicted tissue of origin in which taxane/platinum is standardly used, higher RR and significantly longer PFS and OS were observed compared with pts whose GEP identified a malignancy where taxane/platinum is not standard (Table). Conclusions: In pts with untreated CUP, Carb +Pac +E demonstrated promising antitumor activity. The GEP test identified patients clinically responsive to Carb/Pac/E therapy, and may be useful to select CUP pts for specific antitumor regimens. Clinical trial information: NCT00936702. [Table: see text]

504 Background: CYP2D6 genetic variants and concurrent administration of potent inhibitors (Inh) of the enzyme markedly reduce the plasma concentrations of endoxifen (Jin. JNCI 2005;97:30–9). We have shown that genetic variation in CYP2D6 is associated with a higher risk of relapse in Tam treated patients (pts) (Goetz. JCO 2005;23:9312–18), but the impact of CYP2D6 inhibition on Tam clinical outcome is unknown. Methods: Using the North Central Cancer Treatment Group 89–30–52 adjuvant Tam trial (1989–1994), we evaluated the effect of co-administration of CYP2D6 Inh and CYP2D6*4 genotype on relapse in pts receiving tam alone. Patient charts were reviewed at each randomizing site to record the following known CYP2D6 Inh: fluoxetine (F), paroxetine (P), sertraline (S), cimetidine (C), amiodarone (A), doxepin (D), ticlopidine (T) and haloperidol (H). Poor metabolizers (PM) were defined as pts who were CYP2D6 *4/*4 and/or known to receive a CYP2D6 Inh. Intermediate metabolizers (IM) were defined as WT/*4 without CYP2D6 Inh, and extensive metabolizers (EM) as WT/WT without CYP2D6 Inh. The association between CYP2D6 status and Tam clinical outcome was determined using the log-rank test. Multivariate Cox modeling was performed using traditional prognostic factors. Results: CYP2D6*4 genotype was obtained in 190 of 256 eligible pts, with a CYP2D6*4/*4 genotype frequency of 6.8% as described previously (Goetz JCO 2005). Medication history was available in 171/256 eligible pts and in 143/190 pts with CYP2D6 genotype. Eleven of 173 patients were concomitantly administered these CYP2D6 Inh: F(2), P(2), S(2), H(1), C(4), and D(1). The median duration of use was 2.5 yrs. Twenty-four pts met criteria for PM [13 by *4/*4 genotype and 11 by CYP2D6 Inh (7 WT/WT, 1 WT/*4, and 3 unknown genotype)], 21 for IM, and 102 for EM. Compared with either IM or EM, PMs had significantly worse time to recurrence and disease free survival in both the univariate (HR 2.5, p=0.02; and 2.2, p=0.01) and multivariate (HR 2.7, p=0.01; and HR 2.2 p=.02) analyses utilizing tumor size, nodal status, and tumor grade. Conclusion: Our data indicate that CYP2D6 status can be used to identify pts who should receive tam therapy and tam treated pts should not be co-administered potent CYP2D6 inhibitors. No significant financial relationships to disclose.

Abstract Background: Macrophages are well-established components of the inflammatory repertoire in breast cancer (BC) progression. The role of macrophages in early breast carcinogenesis is less understood. Macrophages adopt microenvironment-driven pro- or anti-inflammatory phenotypes. We evaluated macrophagic subsets in benign breast stroma to evaluate their association with future or concurrent BC. Methods: With institutional approval, FFPE tissues were identified from age-matched subjects, [with index benign breast biopsies and (a subset) interval-matched second benign tissues taken at the time of one of three events: ER+ BC, ER- BC or no cancer event (Controls) (index N=32, 26 and 30 respectively)]. Tissues were immunostained (CD68 or CD163) and stromal CD68+ and CD163+ macrophagic cell densities (MCD) (cells/mm2 stromal fibroadipose area) were digitally quantified. Statistical analysis was performed using Wilcoxon signed-rank tests for univariate analysis and conditional logistic regression models to adjust for BMI. Results: In the index age-matched biopsies (median age 54y), within subjects in all groups, the median CD163+ MCD (54.5; range: 13.9-371.3) was significantly higher than CD68+ MCD (19.6, range: 3.5-78.5) but the two were strongly correlated (r = 0.76); neither correlated with BMI (r&lt; 0.1 each). Across the groups, index benign biopsies of ER- BC had significantly lower CD68+ MCD and CD163+ MCD compared to ER+ BC (CD163: median 47.2 vs. 57.8, p=0.004; and median CD68+ MCD: 15.8 vs. 29.7, p=0.03); and ER- BC had significantly lower CD163+ MCD density than Controls (p=0.02), (all by univariate analysis and after adjusting for BMI). Comparing the index biopsy and second biopsy/excision within subjects, all showed significant increases in CD68+ MCD (median increase 27.9 for ER- BC, 16.1 for ER+ BC, and 17.8 for controls, p&lt;0.01), and only ER-BC showed an increase in CD163+ MCD (median increase 26.6, p=0.01). Thus, in the second biopsies/excisions, the CD163+ MCD and CD68+ MCD were similar across groups (median CD163+ MCD: 69.3, 75.4, and 59.3 and median CD68+ MCD: 47.4, 45.2, and 36.0 for ER+ BC, ER- BC, and controls); however, all groups had decreased CD163+:CD68+ MCD ratios compared to the index biopsies. Conclusion: Benign breast tissues have higher stromal CD163+ MCD than CD68+ MCD irrespective of future BC events and independent of BMI. However, between index and second biopsies/excisions, the stromal CD68+ MCD differentially increased in all groups, lowering CD163+:CD68+ MCD ratios, and likely shifting pro- and anti-inflammatory stromal macrophage functionality. Across groups, ER- BC index biopsies had significantly lower stromal MCD of both types compared to ER+ BC, and were the only group to increase stromal CD163+ MCD between index biopsy and cancer event. These data suggest that ER+ and ER- BC may have different pre-carcinogenic stromal inflammatory microenvironments. Citation Format: Jodi M. Carter, Ethan P. Heinzen, Tanya L. Hoskin, Stacey J. Winham, Derek C. Radisky, Daniel W. Visscher, Amy C. Degnim. Differential stromal macrophagic profiles precede ER+ and ER- breast carcinogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2705.