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Invasive lobular breast cancer (ILC) is an understudied subtype of breast cancer with late recurrence, metastasis to serosal surfaces, such as the peritoneum, and dismal long-term outcome. We previously reported better patient outcome in ER+ breast cancers with high glucocorticoid receptor (GR) expression (Pan et al 2011). Preliminary data suggests more favorable patient outlook in the ILC subtype when tumors express higher levels of GR. The dynamic interaction between a tumor and its microenvironment leads to phenotypic changes in stromal cells and the extracellular matrix to promote growth or invasion of malignant cells. ILC is histologically distinct from invasive ductal carcinoma and is characterized by discohesive tumor cells that grow as “Indian files” due to lack of the cell adhesion molecule E-cadherin. Because of these differences we expect the tumor microenvironment (TME) to be quite unique in ILC compared to other breast cancer subtypes. We hypothesized that differing levels of expression of nuclear receptors by tumor cells would impact cells residing within the stroma, presumably through paracrine signaling. Immunohistochemistry for 25 ILC biopsies revealed a wide variation in GR expression, ranging from completely negative to strongly positive. Our goal was to gain insight into how GR presence or absence in ILC cells might impact gene and protein expression in malignant cells and their TME. We were inspired to examine how crosstalk between the GR-positive or GR-negative cancer cells and their respective tumor microenvironment (TME) differentially impact stromal cell gene expression as well as the immune cell milieu. Bulk expression profiling is inadequate to examine molecular profiles of the tumor and stroma separately. Therefore, we used nanoString GeoMx® high-plex digital spatial profiling of RNA and protein expression in GR-positive and GR-negative primary ILC. We first performed spatial gene expression using the Whole Transcriptome Atlas (WTA) for six primary ILC, two of which were strongly positive for GR, two GR-negative and 2 tumors harboring both GR-positive and GR-negative regions. To examine the tumor cells and the stroma independently we segmented regions into panCK-positive and panCK-negative segments and analyzed those separately. Intriguing differences were observed between GR-positive and negative ILC, for the tumor cells as well as for the TME. Implementation of the Spatial Deconvolution script embedded within the GeoMx® software revealed striking differences in the abundance of immune cells, endothelial cells and fibroblasts in the stroma of GR-positive vs GR-negative ILC. Macrophages and other myeloid subsets were present in significantly higher numbers in the TME of GR-positive compared to GR-negative ILC. The contribution of B- and T-cell subsets as well as endothelial cells and fibroblasts were also notably different. To more precisely define the spatial distribution and abundance of various cell subtypes and their association with tumor cells, we are including protein DSP as an added layer to complement the WTA. We are also performing RNA and protein DSP for an additional ten GR-positive and ten GR-negative ILC. Thus, we will test our initial observations and expand our investigation to generate comprehensive molecular profiles of ILC tumors and stromal cells and acquire a deeper understanding of how GR expression/activation influences the crosstalk between ILC cells and their TME. We anticipate that differences in gene and protein expression will provide clues as to how GR activation affects proliferative and adhesive properties of ILC through modification of the TME. Citation Format: Lynda B. Bennett, Candace Frerich, Sunati Sahoo, Cheryl Lewis, Indu Raman, Min Xu, Guanchun Chen, Suzanne D. Conzen. Digital spatial profiling of RNA and protein in the invasive lobular breast cancer microenvironment [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P2-21-08.

Context.— Neoadjuvant systemic therapy refers to the use of systemic agent(s) for malignancy prior to surgical treatment and has recently emerged as an option for most breast cancer patients eligible for adjuvant systemic therapy. Consequently, treated breast carcinomas have become routine specimens in pathology practices. A standard protocol has not yet been universally adopted for the evaluation and reporting of these specimens. The American Joint Committee on Cancer staging system recognizes the challenges in staging breast carcinomas after neoadjuvant treatment and provides important data points but does not currently provide detailed guidance in estimating the residual tumor burden in the breast and lymph nodes. The Residual Cancer Burden system is the only Web-based system that quantifies treatment response as a continuous variable using residual tumor burden in the breast and the lymph nodes. Objective.— To provide clarifications and guidance for evaluation and reporting of postneoadjuvant breast specimens, discuss issues with the current staging and reporting systems, and provide specific suggestions for future modifications to the American Joint Committee on Cancer system and the Residual Cancer Burden calculator. Data Sources.— English-language literature on the subject and the data from the I-SPY 2, a multicenter, adaptive randomization phase 2 neoadjuvant platform trial for early-stage, high-risk breast cancer patients. Conclusions.— This article highlights challenges in the pathologic evaluation and reporting of treated breast carcinomas and provides recommendations and clarifications for pathologists and clinicians. It also provides specific recommendations for staging and discusses future directions.

AimsTouch preparation (TP) and frozen section (FS) are the two methods routinely used in the intraoperative evaluation (IOE) of sentinel lymph nodes (SLNs) to detect metastases in patients with breast cancer. Both methods are extremely sensitive and specific in the primary surgery (non-neoadjuvant systemic therapy (non-NST)) setting. Since NST introduces unique challenges in the IOE of SLNs, the aim was to determine the accuracy of TP and FS in the IOE of SLNs in the NST setting and compare the results with the non-NST setting and to examine factors that contribute to any differences.MethodsWe analysed 871 SLNs from 232 patients (615 SLNs from NST and 256 SLNs from non-NST settings) between 2016 through 2019.ResultsIn the NST group, TP alone (n=366) had a sensitivity of 45.7% and specificity of 99.7%; FS alone (n=90) had a sensitivity of 83.3% and specificity of 100%. When both TP and FS (n=135) were used, the sensitivity was 80.3% and the specificity was 98.6%.In the non-NST group, TP alone (n=193) had a sensitivity of 66.7% and specificity of 100%; FS alone (n=22) had a sensitivity and specificity of 100%; and combined TP and FS (n=34) had a sensitivity and specificity of 100% and 96%, respectively.ConclusionsEvaluating SLNs intraoperatively in the NST setting can be challenging secondary to therapy-related changes. In the NST setting, FS has higher sensitivity and specificity compared with TP for the IOE of SLNs and should be the preferred method.

Background: I-SPY 2 is a neoadjuvant platform trial open to patients with locally advanced, molecular high-risk breast cancer. In a concerted pursuit of mid-therapy response biomarkers, we evaluated inter-regimen biopsies, to identify patients who may be candidates for treatment de-escalation. In a pilot study, we observed that absence of carcinoma in an inter-regimen biopsy may predict pathologic complete response (pCR). In this expanded study of 100 participants, we sought to confirm that finding and assess pathologic features of the inter-regimen biopsy as predictors of tumor response to neoadjuvant therapy. Methods: Digital H&E images of 100 inter-regimen (12 week) image-guided breast biopsies +/- ancillary immunohistochemistry (p63 and/or cytokeratin) were reviewed by 9 I-SPY affiliated pathologists to record 1) tumor bed and 2) presence/absence of residual invasive carcinoma (IC) (with tumor cellularity scored as 0-100%). The data set included 393 cores (mean 3.9 (2-4) cores/biopsy). Fisher’s exact t-test was used for association of presence/absence of IC with pCR, and tumoral hormone receptor (HR) and HER2 status. Association between biopsy tumor cellularity and residual cancer burden (RCB) indices used Pearson’s correlation. Results: In the biopsy set, 84 (84%) had ≥80% inter-observer diagnostic agreement on both 1) presence of tumor bed and 2) presence/absence of IC (53 IC+ /31 IC-). IC+/IC- biopsies had equal numbers of evaluable tissue cores. The primary tumors were 63% HR+/37% HR-. The presence of IC in the biopsy correlated with tumoral HR/HER2 status (p=0.0014: 74%: HR+HER2-; 62%: TN; 60%: HR+HER2+; 10%: HR-HER2+). Of 31 patients with IC- biopsies, 25 (80%) went on to pCR, whereas only 7/53 (13%) of patients with IC+ biopsies had pCR, conferring an odds ratio for pCR of 26, Fisher p=7.5E-10. Overall, IC- biopsies had a positive predictive value (PPV) for pCR of 81%, with a PPV for HR- tumors of 94% vs. 67% for HR+ tumors (Table 1). In the 6 IC- biopsies from patients with non-pCR (“false-negatives”), most were HR+ (5/6, Table 1), and tumor bed size in the resection specimen was smaller than for IC+ biopsies with non-pCR: 276 mm2 (0.4-1000 mm2) vs. 1166 mm2 (1-11960 mm2). In contrast, the 46/53 IC+ biopsies in patients with non-pCR had a PPV for predicting non-pCR of 86%, (PPV for HR+ tumors: 94% vs. PPV for HR- tumors: 66%. Tumor cellularity in the biopsy (mean 37%, [2.5-93%]) did not correlate with RCB index (p=0.57) or RCB breast-only index (p = 0.17) at resection. Conclusion: In this 100 biopsy set from the I-SPY2 trial, the absence of residual carcinoma in inter-regimen biopsies was highly predictive of pCR, particularly for HR- tumors. The “false-negative” biopsies (IC-/non-pCR) were predominantly HR+ tumors with small residual tumor beds at resection. Conversely, the presence of carcinoma in inter-regimen biopsies was highly predictive of non-pCR, particularly for HR+ tumors. These data demonstrate the utility, and the limitations, of the inter-regimen biopsy as one tool to identify patients who may benefit from therapeutic de-escalation. Table 1: PPV for pCR/non-PCR by Inter-regimen Biopsy StatusInter-regimen biopsy with or without Invasive carcinoma (IC+/-)pCRnon-pCRPPV (Sensitivity) for pCR(IC- Biopsies)PPV (Sensitivity) for non-pCR(IC+ biopsies)IC- biopsiesAll25681% (78%)-HR+10567% (83%)-HR-15194% (75%)-IC+ biopsiesAll746-86% (88%)HR+236-94% (88%)HR-510-66% (91%) Citation Format: Jodi M Carter, Molly E Klein, Sara J Venters, Kimmie Rabe, I Tolgay Ocal, Kamaljeet Singh, Denise M Wolf, Sunati Sahoo, Shuko Harada, Laila Khazai, Malini Harigopal, Alexander D Borowsky, Gregor Krings, Ronald Balassanian, Yunn-Yi Chen, Kimberley Cole, Sonal Shad, Amy Delson, Lamorna Brown-Swigart, I-SPY 2 TRIAL Consortium, Laura Esserman, Laura van ‘t Veer, W Fraser Symmans. Pathologic features of the inter-regimen biopsy predict response to neoadjuvant therapy in the I-SPY 2 TRIAL [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS4-09.

Background: The I-SPY 2 TRIAL, open to patients with locally advanced, molecular high-risk breast cancer, aims to bring each patient to pathologic complete response (pCR) with a minimum of toxicity. Here we test the hypothesis that imaging (MR volume predictors) combined with core biopsy may be used to accurately select candidates who show early response and provide an option of treatment de-escalation at mid-therapy (12 weeks). Methods: Of 100 I-SPY 2 patients with pathologist-assessed core biopsies at the inter-regimen time point (~12 weeks through treatment) and pCR data, 87 also had serial MR images and were considered in this study. Eleven I-SPY 2 TRIAL pathologists independently provided a digital assessment of the presence or absence of residual invasive cancer from H&E stained, and any requested ancillary IHC, images from imaging-guided core biopsies. Pathology predicts pCR if there is a consensus of no invasive residual disease. We generated predictions for all (55) unique pairs over the 11 pathologists, where pCR is predicted if both pathologists find no invasive cells. MRI pCR prediction models were previously developed on an independent dataset of ~990 I-SPY 2 patients, and applied to this cohort. Volume-based prediction models were previously optimized within each subtype and predicted probability thresholds were selected over a range of positive predictive value (PPV). In this study, MR predicts pCR (positive test) if the predicted probability is above a threshold that yields a given PPV value. For each pathologist pair, we combined pathology-based and MR-based predictors into a predictive-RCB (pre-RCB); and pre-RCB predicts a patient as pCR (RCB0) if both MR and pathology predicts pCR. Predictive performance is assessed by calculating the mean and range of PPV and sensitivity.Results: 39% (34/87) of the patients in this study achieved pCR. Over all pairs of pathologists, on average 80% of pathology-only predicted pCRs were true pCRs (mean PPV = 80% [range: 69-92%]), and 74% of patients who achieved pCR were predicted pCR by pathology alone (mean sensitivity = 74% [65-82%]). We assessed combinations with MR probability thresholds at PPV levels 50%-70%; and observed the best balance of PPV and sensitivity for the pre-RCB when MR thresholds were set at 50% PPV level. At this threshold setting, the pre-RCB achieved a PPV = 92% [83-100%], meaning on average 92% of predicted pCRs were true pCRs, and this improvement in positive predictive performance over pathology alone is achieved with a lower but still-reasonable 53% sensitivity [33-62%]. Conclusion: Pre-RCB, which predicts a patient as pCR if both MR and inter-regimen pathology predicts pCR, provides clinically actionable accuracy for treatment de-escalation for early responders (PPV>90%). Adding a final MR review at the time of early surgery may further improve performance. Resulting from data presented in this abstract, the pre-RCB algorithm, including the final MR review, has been operationalized and will be used prospectively to identify patients who are highly likely to have already achieved pCR by the inter-regimen timepoint. Citation Format: Sara J Venters, Wen Li, Denise M Wolf, Jodi M Carter, Molly E Klein, Kamaljeet Singh, Kimmie Rabe, I Tolgay Ocal, David Newitt, Christina Yau, Natsuko Onishi, Jessica Gibbs, Sunati Sahoo, Shuko Harada, Laila Khazai, Malini Harigopal, Alexander D Borowsky, Gregor Krings, Ronald Balassanian, Yunn-Yi Chen, Kimberley Cole, Sonal Shad, Barbara LeStage, Amy Delson, Sandra Finestone, Lamorna Brown-Swigart, I-SPY 2 Imaging Working Group, I-SPY 2 TRIAL Consortium, Laura Esserman, Laura van ‘t Veer, W Fraser Symmans, Nola M Hylton. Serial MRI and pathology combined to select candidates for therapy de-escalation in the I-SPY 2 TRIAL [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS4-10.

Invasive lobular breast cancer (ILC) is an understudied breast cancer subtype with late recurrence, metastasis to serosal surfaces including the peritoneum, and dismal long-term outcome. The interaction between a tumor and its microenvironment leads to phenotypic changes in stromal cells and the ECM that promote proliferation and invasion of malignant cells. ILC is histologically distinct from invasive ductal carcinoma, characterized by discohesive E-cadherin-negative tumor cells that grow in single file. We expect the TME to be unique in ILC. We hypothesized that differing levels of nuclear receptor expression in tumor cells would impact stromal cell composition, presumably through paracrine signaling. There is a range of glucocorticoid receptor (GR) expression in ILC. We sought to determine how crosstalk between GR+ or GR- ILC cells and their respective TME differentially impact stromal cell gene and protein expression, as well as the immune cell milieu. To profile the tumor and stroma separately, we performed nanoString GeoMx digital spatial profiling (DSP) of RNA and protein expression in GR-positive (+) and GR-negative (-) primary ILC. We performed RNA DSP for 12 primary ILCs: 5 strongly positive for GR, 5 GR- and 2 tumors with mixed GR expression. We also completed DSP of proteins on adjacent tissue sections for a total of 87 proteins. To profile tumor and TME independently we segmented regions into PanCK+ tumor and PanCK- stroma. RNA expression analyses revealed striking differences between GR+ and GR- ILC in both tumor cells and TME. Pathway analyses showed enrichment of genes encoding eukaryotic translation initiation and control factors and ribosomal proteins in GR+ tumor cells, as well as downregulation of immune response and antigen presentation pathways. In the TME of the GR- tumors, we saw significantly higher expression of collagen biosynthesis and extracellular matrix genes. Using a spatial deconvolution tool, we observed gene expression indicating markedly higher abundance of macrophages and other myeloid subsets, endothelial cells and fibroblasts in GR+ ILC stroma. There was also evidence for more regulatory T-cells, suggestive of an immune suppressive microenvironment in GR+ ILC. Protein DSP revealed upregulation of proteins involved in DNA repair and cell survival, and reduction of some immune checkpoint proteins in the GR+ tumor cells. In the TME of GR+ ILC, we observed more Ki-67 and ER-alpha as well as proteins expressed on T-regs, as well as downregulation of cytotoxic proteins such as granzyme A. We conclude that in-depth examination of molecular profiles in both the tumor cells and the TME of ILCs will give insight into how tumor GR expression/activation influences the crosstalk between ILC cells and their microenvironment. As expected, we observed heterogeneity between patients, but DSP will allow in-depth characterization of individual tumors at the molecular level. Citation Format: Lynda B. Bennett, Sunati Sahoo, Cheryl M. Lewis, Indu Raman, Candace Frerich, Guanchun Chen, Min Xu, Suzanne D. Conzen. Digital spatial profiling of RNA and protein in the tumor microenvironment of invasive lobular breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2360.

Background: Patients who achieve a pathological complete response (pCR- defined as no invasive cancer) after neoadjuvant chemotherapy (NAC) for breast cancer (BC) have improved outcomes, but there is still controversy about the significance of residual ductal carcinoma in situ (DCIS) on local recurrence rate (LRR). The I-SPY 2 TRIAL is an adaptive neoadjuvant platform trial evaluating novel experimental regimens in comparison to standard chemotherapy in women with high-risk breast cancer. The purpose of this study is to determine if residual DCIS after NAC in early BC affects LRR in patients with or without residual invasive disease in the I-SPY 2 TRIAL. Methods: 933 I-SPY 2 patients with residual cancer burden (RCB) and follow-up data were included in this analysis. Residual DCIS was defined as any carcinoma in situ > 0% on RCB evaluation. Local recurrence was defined as recurrence in breast, chest wall or locoregional nodes and/or skin and subcutaneous tissue. We stratified our cohort into four groups: those without residual invasive disease (defined as RCB0) ± residual DCIS, and those with residual invasive disease (RCB>0) ± residual DCIS. We estimated LRR within each group using the Kaplan Meier method; and used Cox proportional hazards models to assess LRR differences between groups, with: patients with no residual disease (invasive or in situ) as reference group. Results: Among 933 patients assessed, median follow up time was 3.9 years. RCB 0 status was achieved in 337 patients (36%). Of these, 267 (29%) had no residual DCIS, which represents our reference group, and 70 (7%) had residual DCIS. Among 596 (64%) patients who had RCB>0, 296 (32%) had residual DCIS. For patients with RCB0 without DCIS and RCB0 with DCIS, the LRR at 3 years were similar: 2% vs 3% respectively (Hazard ratio: 1.29 [0.26-6.39]). Results were also similar in the RCB>0 group, with a LRR of 10% at 3 years in those without residual DCIS, and 11% in those with residual DCIS. Both RCB>0 groups had significantly higher LRR when compared to the patients with RCB0 without DCIS (Hazard ratio: 5.25 [2.20-12.5]) and HR 5.85 [2.47-13.9] respectively). Conclusion: There was no association between residual DCIS and LRR after neoadjuvant chemotherapy, regardless of resolution of invasive disease. Further work is needed to determine whether residual DCIS should drive locoregional therapy decisions after neoadjuvant chemotherapy for invasive breast cancer. Citation Format: Marie Osdoit, Christina Yau, W. Fraser Symmans, Judy C. Boughey, Smita M. Asare, Ron Balassanian, Jodi M. Carter, Yunn-Yi Chen, Kimberly Cole, Laila Khazai, Molly Klein, Dina Kokh, Gregor Krings, Sunati Sahoo, Gretchen Ahrendt, Akiko Chiba, Cheryl Ewing, Constantine Godellas, Nora Jaskowiak, Brigid Killelea, Helen Krontiras, Rachael Lancaster, Julie Lang, M. Catherine Lee, Arpana Naik, Roshni Rao, Julia Tchou, Shannon Tierney, Eleni Tousimis, Tod Tuttle, Anne Wallace, I-SPY 2 TRIAL Consortium, Bev Parker, Laura J. Esserman, Rita A. Mukhtar. The impact of residual ductal carcinoma in situ on breast cancer recurrence in the neoadjuvant I-SPY2 TRIAL [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 P3-08-16.

Background: Achieving a pathologic complete response (pCR) has been shown on the patient level to predict excellent long-term event-free survival outcomes. Residual cancer burden (RCB) quantifies the extent of residual disease for patients who did not achieve pCR. A high proportion of metastatic events to the central nervous system (CNS), a known chemotherapy sanctuary site, was previously observed among the small number of relapses in patients achieving a pCR (Symmans et al 2017), raising the possibility that these CNS events may be independent of response in the breast. I-SPY2 is an adaptively randomized, phase II, platform trial that evaluates new drugs and combinations in the neoadjuvant setting for women with high-risk primary breast cancer. In this study, we evaluated the type and sites of recurrences by RCB classes in the I-SPY 2 TRIAL. Methods: I-SPY 2 patients enrolled prior to 11/2016 across 9 experimental and control arms, with available RCB and event-free survival (EFS) data were included in this analysis. The median follow-up is 3.8 years. We summarized the EFS event type, further sub-dividing the distant recurrence events by their site of relapse (CNS-only, CNS and other sites, Non-CNS). We estimated the overall and site-specific distant recurrence incidence in each RCB class at 3 years using a competing risk (Fine-Gray) model. In addition, we assessed the association between RCB and distant recurrence free survival including all distant recurrences (DRFS), as well as excluding the CNS-only recurrences (non-CNS DRFS) using a Cox model. Our statistics do not adjust for multiplicities beyond variables evaluated in this study. Results: Among 938 subjects, there were 180 EFS events, including 28 (16%) local recurrences (without distant recurrence and/or death) and 152 DRFS events. Among the DRFS events, 25 patients died without a distant recurrence. 127 experienced distant recurrences, including 22 (17.3%) with CNS-only, 16 (12.6%) with CNS and other sites, 87 (68.5%) with non-CNS distant recurrence; 2 (1.6%) patients had missing recurrence site information. Incidence of CNS-only recurrences are low and are similar across RCB classes (pCR/RCB-0 (n=338): 1%, RCB-I (n=129): 3%, RCB-II (n=328): 2%, RCB-III (n=143): 2% at 3 years). In contrast, the incidence of non-CNS recurrences increase with increasing RCB (RCB-0: 2%, RCB-I: 4%, RCB-II: 11%, RCB-III: 19% at 3 years). DRFS of RCB-I patients do not significantly differ from those achieving a pCR/RCB-0 (DRFS at 3 years: 92% vs. 95%, hazard ratio: 1.77 (0.87-3.63)); the small numerical difference is further reduced when the CNS-only recurrences are excluded (non-CNS DRFS at 3 years: 95% vs. 96%, hazard ratio: 1.48 (0.61-3.58)). CNS recurrences among DRFS events are proportionally higher within the pCR (5/16 (31%)) and RCB-I (5/12 (42%)) than in the RCB-II (8/57 (14%)) and RCB-III (4/42 (9%)) groups largely because of the relative low frequency of non-CNS recurrence events. Conclusions: In our high-risk I-SPY 2 cohort, CNS-only recurrences are uncommon but appear similar across RCB groups, independent of response, suggesting that the CNS is a treatment sanctuary site. In contrast, non-CNS recurrence rates increase as RCB increases. These findings, if confirmed, support the use of RCB to identify patients with excellent outcomes beyond those achieving a pCR; and suggest that inclusion of CNS only recurrences as an outcome event may impact the association between neoadjuvant therapy response and long-term outcome. Citation Format: Christina Yau, Angela DeMichele, W. Fraser Symmans, Lajos Pusztai, Douglas Yee, Amy S. Clark, Christos Hatzis, Jeffrey B. Matthews, Jodi Carter, Yunn-Yi Chen, Kimberly Cole, Laila Khazai, Molly Klein, Dina Kokh, Gregor Krings, Sunati Sahoo, Kathy S. Albain, A. Jo Chien, Kirsten K. Edmiston, Anthony D. Elias, Erin D. Ellis, David M. Euhus, Heather S. Han, Claudine Isaacs, Qamar J. Khan, Julie E. Lang, Janice Lu, Jane L. Meisel, Zaha Mitri, Rita Nanda, Donald W. Northfelt, Tara Sanft, Erica Stringer-Reasor, Rebecca K. Viscusi, Anne M. Wallace, Rachel Yung, Nola M. Hylton, Judy C. Boughey, Michelle E. Melisko, Jane Perlmutter, Hope S. Rugo, Richard Schwab, Laura J. van' t Veer, Donald A. Berry, Laura J. Esserman. Site of recurrence after neoadjuvant therapy: Clues to biology and impact on endpoints [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 P2-20-02.

Background: The I-SPY 2 TRIAL enrolls women with locally advanced, molecular high-risk breast cancer. An integrated Residual Cancer Burden (iRCB), based on MRI volume change through treatment, is used to predict pathologic complete response (pCR) in the randomization/evaluation Bayesian engine. With the goal of effective de-escalation of treatment for patients exhibiting an early response, biomarkers are being assessed for their ability to predict pCR, alone or with MR data, during treatment. Here, we present the results of a pilot study to examine if invasive tumor cellularity in mid-treatment tissue core biopsies predicts pCR in a 40-patient cohort of I-SPY 2 patients. Other pathologic variables evaluated include Ki67, tumoral histologic features, and stromal tumor-infiltrating lymphocytes (sTILs). Methods: I-SPY 2 TRIAL pathologists (N=4) were provided images of H&E-stained and Ki67 IHC- labelled (DAKO/Agilent, clone MIB-1) core biopsy sections from 40 patients at the inter-regimen time point, ~12-weeks into treatment. Of the 40 patients, 35 had 4 cores, 3 had 3 cores, and 2 had 2 cores assessed. In total, images from 153 cores were evaluated. For each core, pathologists were asked to score the % area occupied by tumor bed (treatment changes and/or residual cancer), % of viable invasive tumor (0-100%) within tumor bed (with Nottingham grading, % Ki67 labelled, and % sTILs, using standardized guidelines). As decided by the pathologist group, only cores with identified tumor bed were included in the initial analysis. Concordance between pathologists was assessed for all scored criteria, using % agreement for dichotomous variables, and Pearson correlation (r)/standard deviation (sd) for continuous variables. The maximum and average cellularity recorded over all cores/patient, averaged over all pathologists, were analyzed for association with pCR using t- tests (significance threshold: p Results: Pathologist were in general agreement about the presence or absence of tumor bed, with greater than 82% agreement between any two (83-96%), and an overall agreement of 77%. For scoring the % of the tumor bed involved by invasive cancer, correlations between pairs of pathologists ranged from 0.79-0.95 (mean(r)=0.87, sd=5%), and agreement on a binary presence/absence of invasive cancer was 78%. Both the mean (t-test: p=7.59E-05) and maximum (t-test: p=0.0012) %invasive tumor at 12 weeks, scored as an average over all pathologists, were significantly higher in patients who did not achieve pCR than in responders. We also treated %invasive cellularity as a dichotomous variable (present/absent). 90% (9/10) of patients scored by all pathologists as 0% invasive tumor cells (absent) achieved a pCR, vs only 20% (6/30) of patients scored as >0% invasive cellularity by one or more pathologists (present) (OR=32, Fisher p=0.0005); yielding a positive predictive value for pCR of 0.9. Ki67 and sTILS at 12 weeks were fairly concordant across pathologists ((r,sd)=(0.92, 8.45%) and (0.82,5.5%), respectively), but did not associate with response (p>0.05 for pCR, RCB01, or RCB index). Tumor histologic grade at 12 weeks, assessed in 29/30 patients with non-zero cellularity, trended toward association (Fisher p=0.078): 44% (4/9) with Grade 3 went on to have a pCR, vs. 15% (2/13) with Grade 2 and 0 with Grade 1. These data demonstrate the utility of invasive tumor cellularity as a predictor of pCR in a clinical setting. Conclusion: In this pilot study we demonstrate that the absence of invasive cancer cells within identified tumor bed in mid-treatment core biopsy samples is highly predictive of pCR. Citation Format: Sara J Venters, Denise M Wolf, Lamorna Brown-Swigart, Christina Yau, Amy L Delson, Bev Parker, Ron Balassanian, Jodi Carter, Yunn-Yi Chen, Kimberly Cole, Laila Khazai, Molly Klein, Dina Kokh, Gregor Krings, Sunati Sahoo, Jane Wei, I-SPY 2 TRIAL Consortium, Laura J Esserman, Laura J van't Veer, W Fraser Symmans. Assessing biomarkers to inform treatment de-escalation: Mid-treatment biopsy cellularity predicts pCR in the I-SPY 2 Trial [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 P6-10-02.

Background: Anti-HER2 therapy is a standard treatment for HER2-positive tumors, however, the response to targeted therapy varies with a pathologic complete response rate of approximately 40%. Additionally, the relationship between the level of HER2 gene amplification and the clinical outcome is unclear. Purpose: The aim of this study was to determine clinico-pathologic predictors of pathologic complete response after neoadjuvant targeted therapy in HER2 positive breast cancers. Materials and Methods: We performed a retrospective review of HER2-positive breast cancer patients treated with Neoadjuvant Systemic Therapy (NST). PCR was defined as no residual invasive carcinoma in the breast and axillary lymph nodes (ypT0/is, pN0). We divided our cohort into two groups: patient with pathologic complete response (pCR) and patients without complete responses (non-pCR). We compared the HER2 copy number, HER2/CEP-17 ratio, and clinical and pathologic features using univariate and multivariate analyses. Results: A total of 163 patients with HER2 gene amplification (copy number ≥4) or HER2/CEP17 ratio ≥ 2.0 who received neoadjuvant chemotherapy along with Trastuzumab or Trastuzumab-Pertuzumab therapy were included in the study. Only 33.3% of patients achieved pCR. Patients who achieved pCR had tumors with higher HER2 copy numbers (median: 19.1, range: 4.5 - 45.6), compared to those who did not achieve pCR (median:15.22, range: 4.12 - 35.0, p=0.003). The HER2/CEP17 ratio was higher in the pCR group (median 7.43, range 2.1 - 17.94) compared with non-pCR group (median 6.1, range 1.72 - 12.6, p= 0.32). A receiver operating characteristic curve (ROC) to identify the optimal cutoff point to predict pCR revealed the area under the curve for HER2 copy number and HER2/CEP17 ratio to be 0.639 and 0.600, respectively. Univariate analysis showed significant correlation of pCR with HER2 copy number of ≥14 (p=0.003) and an HER2/CEP17 ratio of 4 or greater (p=0.044), and Ki-67 ≥ 40 % (p=0.002). Consistent with previous studies, tumors with negative hormone receptor (HR) statuses had higher pCR rates (p5cm), were not associated with pCR. In our study, all patients with who negative lymph nodes, achieved pCR. In multivariate regression analysis, only HR negativity (OR=2.60, p=0.008), Ki-67 ≥ 40 % (p=0.015 OR=2.625) and HER 2 copy number ≥14 (p=0.009 OR=2.689) remained as a significant factors associated with pCR. Conclusion: Patients with PCR tend to have higher HER/CEP-17 ratios, smaller tumors and negative nodal statuses. Based on our result, HER2 copy number, negative HR status and high KiI-67 are one of the strongest predictors of pCR in HER2 positive tumors. Citation Format: Mariela Huerta Rosario, Sunati Sahoo. Predictors of pathologic complete response in HER2 positive breast cancer patients treated with neoadjuvant targeted therapy [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 P3-08-63.

Background: Previous studies have independently validated the prognostic relevance of residual cancer burden (RCB) after neoadjuvant chemotherapy. We used results from several independent cohorts in a pooled patient-level analysis to evaluate the relationship of RCB with long-term prognosis across different phenotypic subtypes of breast cancer, to assess generalisability in a broad range of practice settings. Methods: In this pooled analysis, 12 institutes and trials in Europe and the USA were identified by personal communications with site investigators. We obtained participant-level RCB results, and data on clinical and pathological stage, tumour subtype and grade, and treatment and follow-up in November, 2019, from patients (aged ≥18 years) with primary stage I–III breast cancer treated with neoadjuvant chemotherapy followed by surgery. We assessed the association between the continuous RCB score and the primary study outcome, event-free survival, using mixed-effects Cox models with the incorporation of random RCB and cohort effects to account for between-study heterogeneity, and stratification to account for differences in baseline hazard across cancer subtypes defined by hormone receptor status and HER2 status. The association was further evaluated within each breast cancer subtype in multivariable analyses incorporating random RCB and cohort effects and adjustments for age and pretreatment clinical T category, nodal status, and tumour grade. Kaplan-Meier estimates of event-free survival at 3, 5, and 10 years were computed for each RCB class within each subtype. Findings: We analysed participant-level data from 5161 patients treated with neoadjuvant chemotherapy between Sept 12, 1994, and Feb 11, 2019. Median age was 49 years (IQR 20–80). 1164 event-free survival events occurred during follow-up (median follow-up 56 months [IQR 0–186]). RCB score was prognostic within each breast cancer subtype, with higher RCB score significantly associated with worse event-free survival. The univariable hazard ratio (HR) associated with one unit increase in RCB ranged from 1·55 (95% CI 1·41–1·71) for hormone receptor-positive, HER2-negative patients to 2·16 (1·79–2·61) for the hormone receptor-negative, HER2-positive group (with or without HER2-targeted therapy; p

ImportancePathologic complete response (pCR) after neoadjuvant chemotherapy (NAC) in breast cancer strongly correlates with overall survival and has become the standard end point in neoadjuvant trials. However, there is controversy regarding whether the definition of pCR should exclude or permit the presence of residual ductal carcinoma in situ (DCIS).ObjectiveTo examine the association of residual DCIS in surgical specimens after neoadjuvant chemotherapy for breast cancer with survival end points to inform standards for the assessment of pathologic complete response.Design, Setting, and ParticipantsThe study team analyzed the association of residual DCIS after NAC with 3-year event-free survival (EFS), distant recurrence-free survival (DRFS), and local-regional recurrence (LRR) in the I-SPY2 trial, an adaptive neoadjuvant platform trial for patients with breast cancer at high risk of recurrence. This is a retrospective analysis of clinical specimens and data from the ongoing I-SPY2 adaptive platform trial of novel therapeutics on a background of standard of care for early breast cancer. I-SPY2 participants are adult women diagnosed with stage II/III breast cancer at high risk of recurrence.InterventionsParticipants were randomized to receive taxane and anthracycline-based neoadjuvant therapy with or without 1 of 10 investigational agents, followed by definitive surgery.Main Outcomes and MeasuresThe presence of DCIS and EFS, DRFS, and LRR.ResultsThe study team identified 933 I-SPY2 participants (aged 24 to 77 years) with complete pathology and follow-up data. Median follow-up time was 3.9 years; 337 participants (36%) had no residual invasive disease (residual cancer burden 0, or pCR). Of the 337 participants with pCR, 70 (21%) had residual DCIS, which varied significantly by tumor-receptor subtype; residual DCIS was present in 8.5% of triple negative tumors, 15.6% of hormone-receptor positive tumors, and 36.6% of ERBB2-positive tumors. Among those participants with pCR, there was no significant difference in EFS, DRFS, or LRR based on presence or absence of residual DCIS.Conclusions and RelevanceThe analysis supports the definition of pCR as the absence of invasive disease after NAC regardless of the presence or absence of DCIS.Trial RegistrationClinicalTrials.gov Identifier NCT01042379.

Importance Residual cancer burden (RCB) distributions may improve the interpretation of efficacy in neoadjuvant breast cancer trials. Objective To compare RCB distributions between randomized control and investigational treatments within subtypes of breast cancer and explore the relationship with survival. Design, Setting, and Participants The I-SPY2 is a multicenter, platform adaptive, randomized clinical trial in the US that compares, by subtype, investigational agents in combination with chemotherapy vs chemotherapy alone in adult women with stage 2/3 breast cancer at high risk of early recurrence. Investigational treatments graduated in a prespecified subtype if there was 85% or greater predicted probability of higher rate of pathologic complete response (pCR) in a confirmatory, 300-patient, 1:1 randomized, neoadjuvant trial in that subtype. Evaluation of a secondary end point was reported from the 10 investigational agents tested in the I-SPY2 trial from March 200 through 2016, and analyzed as of September 9, 2020. The analysis plan included modeling of RCB within subtypes defined by hormone receptor (HR) andERBB2 status and compared control treatments with investigational treatments that graduated and those that did not graduate. Interventions Neoadjuvant paclitaxel plus/minus 1 of several investigational agents for 12 weeks, then 12 weeks of cyclophosphamide/doxorubicin chemotherapy followed by surgery. Main Outcomes and Measures Residual cancer burden (pathological measure of residual disease) and event-free survival (EFS). Results A total of 938 women (mean [SD] age, 49 [11] years; 66 [7%] Asian, 103 [11%] Black, and 750 [80%] White individuals) from the first 10 investigational agents were included, with a median follow-up of 52 months (IQR, 29 months). Event-free survival worsened significantly per unit of RCB in every subtype of breast cancer (HR-positive/ERBB2-negative: hazard ratio [HZR], 1.75; 95% CI, 1.45-2.16; HR-positive/ERBB2-positive: HZR, 1.55; 95% CI, 1.18-2.05; HR-negative/ERBB2-positive: HZR, 2.39; 95% CI, 1.64-3.49; HR-negative/ERBB2-negative: HZR, 1.99; 95% CI, 1.71-2.31). Prognostic information from RCB was similar from treatments that graduated (HZR, 2.00; 95% CI, 1.57-2.55; 254 [27%]), did not graduate (HZR, 1.87; 95% CI, 1.61-2.17; 486 [52%]), or were control (HZR, 1.79; 95% CI, 1.42-2.26; 198 [21%]). Investigational treatments significantly lowered RCB in HR-negative/ERBB2-negative (graduated and nongraduated treatments) andERBB2-positive subtypes (graduated treatments), with improved EFS (HZR, 0.61; 95% CI, 0.41-0.93) in the exploratory analysis. Conclusions and Relevance In this randomized clinical trial, the prognostic significance of RCB was consistent regardless of subtype and treatment. Effective neoadjuvant treatments shifted the distribution of RCB in addition to increasing pCR rate and appeared to improve EFS. Using a standardized quantitative method to measure response advances the interpretation of efficacy. Trial Registration ClinicalTrials.gov Identifier:NCT01042379

A comparative analysis of clinical and pathologic characteristics of HER2 positive breast cancer patients treated with adjuvant versus neoadjuvant antiHer2 therapy: an analysis of 397 cases. Introduction: Currently there are several anti-HER2 therapy options for patients with HER2 positive breast cancer. Surgery as an initial treatment is usually performed in patients with smaller, node negative tumors. Neoadjuvant therapy (NAT) is the treatment of choice for patients with higher stage disease. Detailed analysis of clinical and pathologic characteristics of patients who received adjuvant versus NAT anti-HER2 therapy has not been well elucidated. Objectives: A comparative analysis of clinical and pathologic findings including biomarker expression (ER, PR, HER2 and Ki67) was performed to determine if there were differences in tumor characteristics and clinical outcome in the two groups.Methods:We retrospectively analyzed data on HER2+ breast cancer patients treated with adjuvant and NAT anti-HER2 therapy from 2011 to 2017. Clinical and pathologic parameters including biomarker expression prior to the start of therapy were obtained from the electronic database after IRB approval. In the adjuvant group, patients were treated with initial surgery followed by anti-HER2 therapy plus chemotherapy. In the NAT group, anti-HER2 therapy plus chemotherapy was administered prior to definitive surgery. Types of anti-HER2 therapies and follow-up information were obtained from the electronic medical record. Results:We identified 258 (64.9%) patients who received NAT and 139 (35.0%) received adjuvant anti-HER2 therapy. Table 1.VariablesNeoadjuvant groupAdjuvant groupp-valueAgeBelow 40 years42 (16.2%)11(7.9%)0.02940 years and above Total216 (83.7%) 258128 (92.0%) 139Menopausal statusPremenopausalPostmenopausal Total113 (45.3%)136 (54.6%) 24935 (25.1%)104 (74.8%) 139 Citation Format: Venetia Sarode, Tricia Rood, Yulun Liu, Yisheng Fang, Sunati Sahoo, Yan Peng, Helena Hwang, Marilyn Leitch, Barbara Haley. A comparative analysis of clinical and pathologic characteristics of patients with HER2 positive breast cancer treated with neoadjuvant versus adjuvant anti-HER2 therapy: Analysis of 397 cases [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P1-02-14.

Background Preclinical breast cancer models with acquired HER2 resistance exhibit decreased proliferation with CDK4/6 inhibition in tumors with intact Rb and low p16 levels. Adding cytotoxic agents like T-DM1 enhances the inhibitory CDK4/6 cytostatic effect. Patients and Methods A phase I/Ib 3+3 dose escalation/expansion trial of palbociclib and T-DM1 identified 150 mg on days 5 to 18 as the palbociclib maximal tolerated dose combined with day 1 intravenous T-DM1 in 21-day treatment cycles. Patients were previously treated with trastuzumab and a taxane with no limitation on prior therapy lines, including prior pertuzumab, lapitinib, neratinib, and T-DM1. Median age was 54 years and two-thirds were estrogen receptor positive. Primary objectives included maximum tolerated dose as determined by dose-limiting toxicity, and secondary end points of safety, toxicity, response rate, response duration, and progression-free survival. Results From May 2014 to August 2018, 18 total patients were treated. The median number of cycles was 6.5 (1–22). A maximum tolerated dose was not reached. The most common G3 toxicity of more than 10% incidence was hematologic. Overall response rate (complete response + partial response) was 33% (95% confidence interval, 13%–59%). Median duration of response in responders was not reached and median-progression free survival was 6 months (95% confidence interval, 2.5–11.6). Conclusions The combination of day 1 T-DM1 and days 5 to 18 palbociclib is safe, tolerable, and active in previously treated HER2-positive relapsed patients. Observed hematologic toxicity is manageable. The trial response rate confirms that a CDK 4/6 inhibitor can resensitize HER2-resistant breast cancer.

CDK4/6 inhibitors (CDK4/6i) in combination with antiestrogens have revolutionized the treatment of ER+ metastatic breast cancer (MBC), significantly prolonging survival. However, this combination is not curative, and tumors eventually acquire resistance. Following progression on this combination, patients are left with limited treatment options. A diverse array of mechanisms of resistance to CDK4/6i + antiestrogens have been described. However, laboratory models that capture this heterogeneity of resistance mechanisms are lacking. Patient-derived organoids (PDOs) provide a rapid, robust and reliable platform that recapitulates intra-tumor heterogeneity, partially mimics the cancer microenvironment, and accurately predicts drug response. We aspired to generate a platform of CDK4/6i-resistant breast cancer PDOs to serve as models for understanding acquired resistance to CDK4/6i + antiestrogens and identifying therapies to overcome resistance. We successfully established 16 PDOs out of 32 biopsies (50% efficiency) of metastates from patients with ER+ MBC progressing on CDK4/6i (palbociclib or abemaciclib) + antiestrogens (letrozole or fulvestrant; median response to combination = 9 months). Our collection includes PDOs derived from lobular (n=3) and inflammatory (n=2) breast cancers and reflects racial/ethnic diversity (50% white/not Hispanic; 18.8% Hispanic; 12.5% Black; 12.5% other/unknown). Next-gen sequencing reports were available for 10 patients from which organoids were established, revealing alterations associated with CDK4/6i and/or antiestrogen resistance, including ESR1 (n=2), HER2/ERBB2 (n=2), PTEN (n=2), CCNE1 (n=1), NF1 (n=1), and ARID1A (n=1). Furthermore, one biopsy and its derived organoid lost ER expression, and 5 harbored PIK3CA activating mutations. Thus far, we have performed targeted DNA-sequencing on 7 PDOs and found 13/15 (86.7%) concordance with driver mutations from tumor NGS reports. PDOs established from CDK4/6i-resistant biopsies maintained resistance to palbociclib or abemaciclib ± fulvestrant (500 nM each) in 3D cell viability assays (6 days of treatment). In contrast, control PDOs established from primary ER+ breast cancer surgical samples (n=2) were sensitive to each CDK4/6i ± fulvestrant (median viability for combination=25.6-31.5% for control vs 65.2-80.5% for resistant). GSEA analysis of RNA-seq data from control (n=2) and CDK4/6i-resistant (n=6) PDOs cultured in estrogen-depleted media ± 200 nM palbociclib revealed that palbociclib treatment resulted in downregulation of E2F target and G2M checkpoint signatures in control but not resistant PDOs. Next, we performed a high-throughput screen of 1,000 compounds in 3 resistant PDOs. One PDO showed exquisite sensitivity to G2/M cell cycle checkpoint components, including CDK1, PLK1, Aurora kinase, ATR, Chk1, and Wee1 inhibitors. Finally, treatment of 10 resistant PDOs with the CDK2/4/6 inhibitor PF-06873600 revealed that the CCNE1 (cyclin E1)-amplified PDO was highly sensitive (IC50=130 nM vs >1000 nM), supporting that CCNE1-amplified tumors are vulnerable to CDK2 inhibition. Conclusions: PDOs can be successfully established from ER+ MBC biopsies, maintain the resistant phenotype in culture, retain driver alterations found in tumors from which they were derived, and fail to suppress E2F targets following treatment with CDK4/6i. Therefore, these PDOs represent valuable models to understand and explore diverse mechanisms of CDK4/6i resistance and therapeutic vulnerabilities. Citation Format: Ariella B. Hanker, Sumanta Chatterjee, Yunguan Wang, Dan Ye, Dhivya R. Sudhan, Brian M. Larsen, Lauren C. Smith, Yilin Zhang, Vishal Kandagatla, Kuntal Majmudar, Ezequiel Renzulli, Saurabh Mendiratta, Kimberly Blackwell, Alana L. Welm, Sunati Sahoo, Nisha Unni, Cheryl M. Lewis, Tao Wang, Ameen A. Salahudeen, Carlos L. Arteaga. A platform of CDK4/6 inhibitor-resistant patient-derived breast cancer organoids illuminates mechanisms of resistance and therapeutic vulnerabilities [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PD2-01.

Background Breast cancer metastases to an ipsilateral supraclavicular lymph node is assigned a N3 status in the TNM system and thus classified as stage III disease in the American Joint Commission on Cancer staging manual. Breast cancer metastatic to contralateral axillary lymph node (CAM) without metastases to any other distant organ is currently assigned M1 status (stage IV) instead of N3 (stage III). Patients and Methods We retrospectively reviewed the medical records of breast cancer patients diagnosed with CAM for their clinical presentation, pathologic diagnoses, treatment, and follow-up data. Patients who had distant metastases at the time of CAM diagnosis were excluded from the study. Results We report 12 breast cancer patients who developed CAM but no evidence of metastases in any other distant organ documented with extensive imaging workup. Imaging studies and thorough pathologic evaluation of the prophylactic total mastectomy specimen did not reveal a primary in the breast to account for the metastases in the axillary node. Conclusion Findings of our study as well as previous studies support that lymph node metastases in the contralateral axilla represents a locoregional spread of the tumor from the index breast via lymphatics rather than hematogenous spread. Therefore, isolated CAM in breast cancer patients should not be classified as stage IV disease.

Background: Pathological complete response (pCR) is accepted by FDA as a surrogate endpoint for accelerated approval of targeted agents in combination with chemotherapy based on better long-term outcomes compared to residual disease (Cortazar 2014). Methods: The multi-center, adaptively-randomized I-SPY2 platform trial uses pCR as the primary endpoint to identify investigational agents that will improve outcomes in women with stage 2/3 breast cancer with high risk of early recurrence, across all signatures, based on hormone receptor (HR), HER2, and 70-gene (MammaPrint) status. For patients with HR+ HER2- tumors, only 70-gene (Mammaprint) high-risk patients are enrolled. To date, 1200+ patients have been randomized to one of 14 arms: control (paclitaxel followed by AC); veliparib/carboplatin; neratinib; MK2206; trebananib; trastuzumab/pertuzumab; ado-trastuzumab emtansine/pertuzumab; pembrolizumabx4; ganitumab/metformin; ganetespib; PLX-3397. 7 agents graduated in at least one signature (> 85% probability of success in a 300-patient phase III confirmatory trial); 2 did not graduate; 1 stopped for toxicity, and 3 are enrolling (patritumab/trastuzumab, talazoparib/irinotecan, pembrolizumabx8). Local pathologists were centrally trained using the Residual Cancer Burden (RCB) assessment to ensure uniform evaluation and response classification; RCB 0 = pCR. Results: We evaluated the relationship between pCR and event free (EFS) and distant disease free survival (DDFS) in the first 522 pts (median follow-up:2.5 years). 180 pts achieved pCR (36%) while 338 did not (RCB=1-3). There were 82 EFS and 65 DRFS events. Over the entire group (including all arms), pCR was highly associated with 3-year EFS (p 3-year survival (pCR group)Hazard Ratio OverallOverallHR+/HER2-HER2+TNBCEFS97%0.08 (0.03-0.23)0.14 (0.02-1.04)0 (NA)0.11 (0.03-0.37)DDFS98%0.08 (0.03-0.26)0.17 (0.01-1.23)0 (NA)0.09 (0.02-0.40) Conclusions: The first long-term efficacy results from the I-SPY2 TRIAL demonstrate that achieving pCR is a very strong surrogate endpoint for improved EFS and DDFS in a high-risk population, across all treatment arms, regardless of subtype. I-SPY2 shows substantially lower estimated EFS hazards for patients achieving pCR, compared to the 5 yr EFS hazard ratio for pCR vs not in Cortazar (hazard ratio 0.49), demonstrating important differences between a metaanalysis compared to a platform trial with uniform high-risk eligibility, standardized pathology assessment, and multiple targeted therapies. Our data support the use of pCR as a primary endpoint for accelerated approval of new drugs if EFS is evaluated in the same population. Based on these findings, the I-SPY2 TRIAL will test whether therapy can be deescalated or escalated for individual patients with the goal of achieving pCR for all. Citation Format: Yee D, DeMichele A, Isaacs C, Symmans F, Yau C, Albain KS, Hylton NM, Forero-Torres A, van't Veer LJ, Perlmutter J, Rugo HS, Melisko M, Chen Y-Y, Balassanian R, Krings G, Datnow B, Hasteh F, Tipps A, Weidner N, Zhang H, Tickman R, Thornton S, Ritter J, Amin K, Klein M, Chen B, Keeney G, Ocal T, Feldman M, Klipfel N, Sattar H, Mueller J, Gwin K, Baker G, Kallakury B, Zeck J, Duan X, Ersahin C, Gamez R, Troxell M, Mansoor A, Grasso LeBeau L, Sams S, Wisell J, Wei S, Harada S, Vinh T, Stamatakos MD, Tawfik O, Fan F, Adams A, Rendi M, Minton S, Magliocco A, Sahoo S, Fang Y, Hirst G, Singhrao R, Asare SM, Wallace AM, Chien AJ, Ellis ED, Han HS, Clark AS, Boughey JC, Elias AD, Nanda R, Korde L, Murthy R, Lang J, Northfelt D, Khan Q, Edmiston KK, Viscusi R, Haley B, Kemmer K, Zelnak A, Berry DA, Esserman LJ. Pathological complete response predicts event-free and distant disease-free survival in the I-SPY2 TRIAL [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr GS3-08.

Mammary Paget’s disease is rare and comprises about 0.62% of all breast cancer cases, only 1.65% of which occur in male patients. This case report involves a 76-year-old man who presented to his primary care physician with an itching, scaly, unilateral lesion involving the nipple skin. He underwent wide local excision of the lesion for a diagnosis of Bowen’s disease (squamous cell carcinoma in situ). Histologic examination of the specimen revealed mammary Paget’s disease with ductal carcinoma in situ in the underlying breast tissue. A panel of immunohistochemical stains revealed the Paget cells to be positive for cytokeratin 7, MUC1, GATA3, and androgen receptor and negative for cytokeratins 5/6, p63, SOX10, and MART-1/Melan-A. Paget cells were also negative for estrogen receptor and progesterone receptor, and positive for HER2/neu. However, the underlying ductal carcinoma in situ was positive for both estrogen receptor and progesterone receptor and negative for HER2/neu. This discordance, supported by the current literature, suggests an alternative etiology for Paget’s disease in certain cases that cannot be explained by the well-established epidermotropic and transformative theories of Paget’s disease evolution.

We compared the performance of two Food and Drug Administration-approved HER2 immunohistochemistry (IHC) tests: HercepTest (Dako) and PATHWAY anti-HER2 (4B5) (Ventana).In total, 180 invasive breast carcinomas previously tested by both HercepTest and fluorescent in situ hybridization (FISH) were retested with 4B5. Three pathologists scored the HER2 IHC using the 2013 American Society of Clinical Oncology/College of American Pathologists guidelines. The HER2 IHC results were correlated with FISH.Among 135 equivocal cases by HercepTest, 100 (74.1%) were negative by 4B5. Among 45 positive HercepTest cases 9 (20%) were equivocal by 4B5. Among 135 equivocal HercepTest results, 100 (74.1%) were nonamplified, 18 (13.3%) equivocal, and 17 (12.6%) amplified by FISH. Among the 45 positive results with HercepTest, 2 (4.5%) were nonamplified and 1 (2.2%) was equivocal by FISH. All 37 positive and 3 negative by 4B5 cases were amplified by FISH. The absolute interobserver agreement was high for both tests (Fleiss kappa=0.838 for HercepTest and 0.771 for 4B5).PATHWAY anti-HER2 (4B5) significantly reduced the number of equivocal results that require additional testing. Although HercepTest was positive in a small number of HER2 nonamplified cases, 4B5 failed to detect 3 cases that were interpreted as positive by FISH, all with nonclassic or low levels of amplification.

Breast cancer typically spreads primarily to regional lymph nodes and subsequently to distant sites via hematogenous routes. Occasionally metastasis can occur through lymphangitic spread, usually to the lungs, resulting in lymphangitic carcinomatosis. Lymphangitic spread of several malignancies have been reported at other sites in the body with varying degrees of clinical significance. In this case report, we describe a rare case of lymphangitic spread of invasive lobular carcinoma to the contralateral breast identified on imaging as significant background enhancement without a discrete suspicious mass.

A majority of breast cancer specific deaths in women with ER&alpha, (+) tumors occur due to metastases that are resistant to endocrine therapy. There is a critical need for novel therapeutic approaches to resensitize recurrent ER&alpha, (+) tumors to endocrine therapies. The objective of this study was to elucidate mechanisms of improved effectiveness of combined targeting of ER&alpha, and the nuclear transport protein XPO1 in overcoming endocrine resistance. Selinexor (SEL), an XPO1 antagonist, has been evaluated in multiple late stage clinical trials in patients with relapsed and /or refractory hematological and solid tumor malignancies. Our transcriptomics analysis showed that 4-Hydroxytamoxifen (4-OHT), SEL alone or their combination induced differential Akt signaling- and metabolism-associated gene expression profiles. Western blot analysis in endocrine resistant cell lines and xenograft models validated differential Akt phosphorylation. Using the Seahorse metabolic profiler, we showed that ER&alpha, XPO1 targeting changed the metabolic phenotype of TAM-resistant breast cancer cells from an energetic to a quiescent profile. This finding demonstrated that combined targeting of XPO1 and ER&alpha, rewired the metabolic pathways and shut down both glycolytic and mitochondrial pathways that would eventually lead to autophagy. Remodeling metabolic pathways to regenerate new vulnerabilities in endocrine resistant breast tumors is novel, and given the need for better strategies to improve therapy response in relapsed ER&alpha, (+) tumors, our findings show great promise for uncovering the role that ER&alpha, XPO1 crosstalk plays in reducing cancer recurrences.

Importance Pathologic complete response (pCR) is a known prognostic biomarker for long-term outcomes. The I-SPY2 trial evaluated if the strength of this clinical association persists in the context of a phase 2 neoadjuvant platform trial. Objective To evaluate the association of pCR with event-free survival (EFS) and pCR with distant recurrence–free survival (DRFS) in subpopulations of women with high-risk operable breast cancer treated with standard therapy or one of several novel agents. Design, Setting, and Participants Multicenter platform trial of women with operable clinical stage 2 or 3 breast cancer with no prior surgery or systemic therapy for breast cancer; primary tumors were 2.5 cm or larger. Women with tumors that were ERBB2 negative/hormone receptor (HR) positive with low 70-gene assay score were excluded. Participants were adaptively randomized to one of several different investigational regimens or control therapy within molecular subtypes from March 2010 through 2016. The analysis included participants with follow-up data available as of February 26, 2019. Interventions Standard-of-care neoadjuvant therapy consisting of taxane treatment with or without (as control) one of several investigational agents or combinations followed by doxorubicin and cyclophosphamide. Main Outcomes and Measures Pathologic complete response and 3-year EFS and DRFS. Results Of the 950 participants (median [range] age, 49 [23-77] years), 330 (34.7%) achieved pCR. Three-year EFS and DRFS for patients who achieved pCR were both 95%. Hazard ratios for pCR vs non-pCR were 0.19 for EFS (95% CI, 0.12-0.31) and 0.21 for DRFS (95% CI, 0.13-0.34) and were similar across molecular subtypes, varying from 0.14 to 0.18 for EFS and 0.10 to 0.20 for DRFS. Conclusions and Relevance The 3-year outcomes from the I-SPY2 trial show that, regardless of subtype and/or treatment regimen, including 9 novel therapeutic combinations, achieving pCR after neoadjuvant therapy implies approximately an 80% reduction in recurrence rate. The goal of the I-SPY2 trial is to rapidly identify investigational therapies that may improve pCR when validated in a phase 3 confirmatory trial. Whether pCR is a validated surrogate in the sense that a therapy that improves pCR rate can be assumed to also improve long-term outcome requires further study. Trial Registration ClinicalTrials.gov Identifier:NCT01042379

Background With increased utilization of magnetic resonance imaging (MRI) of the breast in clinical practice, many non-mass lesions are being detected incidentally. Although MRI-guided biopsies are usually performed for these lesions, no clear guidelines exist to determine the management of these lesions. Objective The purpose of this study was to explore the pathological features of incidentally detected non-mass lesions on breast MRI to determine rates of malignancy and to assess the predictive value of kinetic enhancement curves for further characterization of these lesions as benign versus malignant. Methods A retrospective review was performed of all MRI-guided core needle biopsies done at our institution between January 2009 and January 2012. Results 172 consecutively biopsied lesions were identified. 43% of the lesions were non-mass lesions, out of which 74.3% were benign, whereas 14.9% and 10.8% of these lesions had malignant and atypical features, respectively. The most commonly encountered non-mass malignant lesion was ductal carcinoma in situ, whereas the most frequently seen invasive non-mass lesion was invasive lobular carcinoma. Kinetic enhancement curves for these lesions did not demonstrate a statistically significant relationship with their pathologic features. Conclusions Non-mass enhancing lesions incidentally detected on breast MRI carry a significant risk of atypia and malignancy, either in situ or invasive. Hence, it is prudent to biopsy these lesions.

Context.—Breast magnetic resonance imaging (MRI) is now used routinely for high-risk screening and in the evaluation of the extent of disease in newly diagnosed breast cancer patients. Morphologic characteristics and the kinetic pattern largely determine how suspicious a breast lesion is on MRI. Because of its high sensitivity, MRI identifies a large number of suspicious lesions. However, the low to moderate specificity and the additional cost have raised questions regarding its frequent use.Objectives.—To identify the pathologic entities that frequently present as suspicious enhancing lesions and to identify specific MRI characteristics that may be predictive of malignancy.Design.—One hundred seventy-seven MRI-guided biopsies from 152 patients were included in the study. The indication for MRI, MRI features, pathologic findings, and patient demographics were recorded. The MRI findings and the pathology slides were reviewed by a dedicated breast radiologist and breast pathologists.Results.—Seventy-one percent (126 of 177) of MRI-guided breast biopsies were benign, 11% (20 of 177) showed epithelial atypia, and 18% (31 of 177) showed malignancy. The vast majority (84%; 62 of 74) of MRI lesions with persistent kinetics were benign. However, 57% (17 of 30) of lesions with washout kinetics and 65% (62 of 95) of mass lesions were also benign.Conclusions.—Magnetic resonance imaging detects malignancies undetected by other imaging modalities but also detects a wide variety of benign lesions. Benign and malignant lesions identified by MRI share similar morphologic and kinetic features, necessitating biopsy for histologic confirmation.

Background: Triple negative (ER negative, PR negative, Her2 negative) breast cancer is an aggressive cancer that is not likely to respond to endocrine or anti-Her2 therapy. While Her2 positivity is a poor prognostic factor, patients with Her2 positive tumors are eligible for anti-Her2 therapy. Her2 positivity is based on either immunohistochemistry (IHC) or fluorescence in situ hybridization (FISH) result. Her2 IHC scores of 0+ and 1+ are negative, 2+ is equivocal, and 3+ is positive. The Her2 gene is amplified when the Her2/CEP17 ratio is ≥2.0. While most tumors will show IHC/FISH concordance, some tumors may show discordant findings between IHC staining and FISH. Previous studies have shown benefit from anti-Her2 therapy in tumors with discordant IHC/FISH results. The purpose of this study was to determine the frequency of Her2 amplification in triple IHC negative breast cancer to determine whether routine Her2 FISH testing is necessary in this subset of patients. Methods: After Institutional Review Board approval, the pathology databases were searched for cases of triple IHC negative breast cancer from 2003-August 2013. All cases of triple IHC negative with corresponding Her2 FISH testing were included in this study. The results of Her2 FISH were correlated with Her2 IHC. The clinical history and IHC slides for all discordant Her2 cases were reviewed when available. Results: A total of 659 triple IHC negative breast cancer cases from 516 patients were found. The patients’ age at diagnosis ranged from 26-92. The cases were classified using the ratio of Her2/CEP17 per the 2013ASCO/CAP guidelines: non-amplified < 2.0, amplified ≥ 2.0. As the copy number for Her2 was not always available, the 2013 ASCO/CAP criterion for amplification based on the copy number was not used in reclassifying the cases. Based on the ratio, 20 tumors were amplified, 631 tumors were not amplified, and 8 tumors had insufficient tissue for Her2 FISH testing. The Her2/CEP17 ratio for amplified cases ranged from 2.0 to 11.1 and for non-amplified cases from 0.5 to 1.9. Of these 20 patients, 10 received some trastuzumab treatment, 6 did not receive any trastuzumab, and in 4 patients, there was no additional information available regarding treatment with trastuzumab. Of the 10 who received trastuzumab treatment, 2 died of disease (DOD), 26 months and 45 months after their diagnosis. The overall survival of the other patients in this group was 23-106 months. Of the 6 who did not receive trastuzumab, one DOD after 76 months. The other 5 patients have overall survival ranging from 15-93 months. Of the patients with unknown Her2 targeted therapy status, one is deceased, the status of 2 is unknown, and one patient is alive at 110 months. Discussion: Our study showed 3% of triple IHC negative breast cancers to be Her2 FISH amplified. Of the 20 tumors, 19 showed Her2/CEP17 between 2.0 to 4.2 with one case showing Her2/CEP17 of 11.1. Firm conclusions on the efficacy of trastuzumab treatment in this small subset of patients with Her2 amplification but no protein overexpression cannot be drawn. However, the overall survival of those treated and not treated is similar. Based on the small number of cases with Her2 IHC/FISH discordance, a compelling case for routine testing of all Her2 IHC negative tumors cannot be made. Citation Format: Sunati Sahoo, Helena Hwang. Her2 FISH amplification in ER/PR/Her2 IHC negative breast cancer [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P6-08-53.

Neoadjuvant therapy (NAT) originally reserved for the treatment of inflammatory and locally advanced breast cancers is currently offered to women with earlier-stage and operable breast carcinoma. NAT allows more women to be eligible for breast conservation surgery and provides an opportunity to assess the response of carcinomas to therapy. This review focuses on the predictors of therapeutic response in pretreatment tumor, evaluation of post-treatment breast and lymph node specimens and classification systems to evaluate degree of response to NAT.

Atypical lobular hyperplasia (ALH) and lobular carcinoma in situ (LCIS) are lobular intraepithelial neoplasms that are often grouped under the term lobular neoplasia (LN). The pathognomonic feature of LN is discohesion secondary to absence or dysfunction of E-cadherin. LN is divided into classical and variant types and is considered to be both a risk factor and non-obligate precursor of invasive carcinoma. Classical LN is more common, shows indolent behavior, and is managed conservatively. Variant LN includes pleomorphic LCIS and florid LCIS, both of which have examples that would fall under the general category of “LCIS with necrosis.” Variant LN is uncommon and under recognized. Clinical data on variant LN is lacking, however the histology, biomarker profile, molecular alterations, and disease associations suggest a more aggressive disease than classical LN. Currently, there is no clear consensus on management of classical LN when diagnosed on core biopsy, whereas excision is recommended routinely for LN variants.

Invasive lobular carcinoma (ILC) is the second most common invasive breast carcinoma (IBC), comprising approximately 10 % of all IBC. Classical ILC is more likely to be clinically and mammographically occult than IDC and therefore, may present at a later stage. Classical ILC is hormone receptor (HR) positive and HER2 negative. As in IDC, histologic grading of ILC should be performed. High grade ILC is uncommon and may be HR negative and HER2 positive. E-cadherin can aid in diagnosing ILC; however, aberrant staining can occur. Classical ILC exhibits behavior similar to other ER positive tumors, showing late rather than early recurrence, and better response to hormonal therapy compared to chemotherapy. ILC shows a different pattern of metastasis than IDC, involving unusual sites such as the gynecologic and gastrointestinal tracts, and peritoneum.

Invasive ductal carcinoma, not otherwise specified (IDC, NOS), also referred to as invasive breast carcinoma, no special type, (IBC, NST), is the most common invasive carcinoma. It is a diagnosis of exclusion conferred when the tumor cannot be classified as a specific type of breast carcinoma. Consistent with the no special type classification, IDC shows marked heterogeneity in morphology, grade, hormone receptor (HR) and HER2 expression, and prognosis. Invasive tubular and cribriform carcinomas are special types of IBC that are low-grade, invariably HR positive and HER2 negative, and have an excellent prognosis. Microinvasive carcinoma, defined as invasion 1 mm or less in greatest dimension, is usually found in association with carcinoma in situ and shows an overall prognosis similar to ductal carcinoma in situ. Although gene-expression profiling has segregated tumors into different subtypes, classification based on morphologic features is invaluable for some of these special types of breast carcinomas as it determines prognosis and guides treatment options.

Background Prophylactic mastectomies (PM) are performed to reduce the risk of breast cancer. Occasionally an occult carcinoma is found in PM specimens. Given the high morbidity of axillary lymph node dissection (ALND), some perform prophylactic sentinel lymph node biopsy (SLNB). We undertook a study to examine if prophylactic SLNB is indicated in all patients undergoing PM. Methods A retrospective review of all PM between 2004 and 2010 was performed. The stage of tumor on the disease side and the pathologic findings in the prophylactic breast were analyzed. The number of SLN and the frequency of lymph node metastases were evaluated. Results A total of 199 PM on 184 patients were performed: 169 contralateral PM and 30 bilateral PM. Of the 199 PM, 12 had occult carcinomas (6.0%): 10 non-invasive, 1 microinvasive and 1 T1b invasive tumor. 153 of 199 PM specimens had prophylactic SLNB, of which only 2 had a positive SLN that originated from the index side of the breast. Conclusions Although it is not uncommon to find occult carcinomas in the prophylactic breast, it is rare for the occult carcinoma to spread to the lymph nodes. Therefore, routine prophylactic SLNB is not indicated in patients undergoing PM. J. Surg. Oncol. 2012; 105:650–654. © 2011 Wiley Periodicals, Inc.

Majority of breast cancer specific deaths in women with ERα (+) tumor occur due to metastases that are resistant to endocrine therapy. There is a critical need for novel therapeutic approaches to resensitize recurrent ERα (+) tumors to endocrine therapies. The objective of this study was to elucidate mechanisms of improved effectiveness of combined targeting of ERα and XPO1, a nuclear transport protein in overcoming endocrine resistance. Selinexor (SXR), an XPO1 antagonist, has been evaluated in multiple later stage clinical trials in patients with relapsed and /or refractory hematological and solid tumor malignancies. Using Cignalfinder to profile kinase signaling pathways, we found that 4-OH-Tam, SXR or their combination induced differential Akt phosphorylation profiles, changing the localization and activity of the kinase. Since we observed dramatic changes in Akt activity we hypothesized that metabolic profile of breast cancer cells would change in the presence of 4-OH-Tam and SXR. Using Seahorse metabolic profiler and cell viability experiments in limited media conditions we showed that tamoxifen resistant cells were more dependent on mitochondria for energy production. Their glucose and fatty acid dependency decreased in the presence of SXR and cells were more dependent on glutamine as the mitochondrial fuel source. In order to examine metabolic pathways that might result in the observed phenotype we performed transcriptomics and GC/MS whole metabolite profiling and identified aminoacid metabolism pathways to be upregulated when cells were treated with SXR+4-OH-Tam. We demonstrated that combined targeting of XPO1 and ERα rewires metabolic pathways and shuts down both glycolytic and mitochondrial pathways that would eventually lead to autophagy. Remodelling metabolic pathways to regenerate new vulnerabilities in endocrine resistant breast tumors is novel, and given the need for better strategies for improving therapy response of relapsed ERα(+) tumors, our findings show great promise for uncovering the role ERα-XPO1 crosstalk plays in reducing cancer recurrences. Citation Format: Zeynep Madak Erdogan, Eylem Cotul-Kulkoyluoglu, Kinga Wrobel, Sunati Sahoo, Barbara Haley, Yosef Landesman. Combined targeting of estrogen receptor alpha and nuclear transport pathways remodel metabolic pathways to induce autophagy and overcome endocrine resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3733.

Malignant phylloides tumors are exceedingly rare with few cases being reported in pregnancy. We describe the first case ever reported of a malignant phylloides tumor presenting in the first trimester of pregnancy and provide insight into the complexities of management as well as a review of the known literature. An extensive PubMed literature search for “cystosarcoma,” “phylloides,” and “pregnancy” was performed. References of each citation were reviewed. Only six previous cases of phylloides tumor in pregnancy were found, none of which were in the first trimester. Medical records of a patient presenting to our institution at 9 weeks gestation with a malignant phylloides tumor were reviewed. We further provide a review of the current literature of the management of phylloides tumor in pregnancy. A 27-year-old white G2P0SA1 woman with no family history of breast cancer presented with a right breast mass at her first prenatal examination at 9 weeks of pregnancy. Ultrasound confirmed a solid mass measuring 24 mm. Core needle biopsy demonstrated a malignant phylloides tumor. She previously had a fibroadenoma removed from the same breast 7 years previously. The current tumor was excised to clear margins. Histopathological examination revealed a 4-cm fibroepithelial tumor with marked stromal cellularity and a high mitotic count (five to seven mitoses/high-power field), confirming the diagnosis of malignant phylloides tumor. The patient continued her pregnancy without complications. Six other cases of phylloides tumor presenting in pregnancy have been reported in the literature, one of which had bilateral disease. Of these, the average patient age was 32 years (range, 28 to 35 years). The majority of these patients presented in their third trimester (mean, 29 weeks; range, 20 to 36 weeks) and often had large tumors (mean, 15 cm; range, 5 to 21 cm). Four of the seven tumors (57%) required a mastectomy. Previous cases have shown phylloides tumors to present in the third trimester as large masses that require mastectomy. With early detection, malignant phylloides tumors can present in the first trimester of pregnancy at smaller sizes; in these patients, breast-conserving surgery is possible.

Currently radiologists have the option of subcategorizing BI-RADS 4 breast lesions into 4A (low suspicion for malignancy), 4B (intermediate suspicion of malignancy), and 4C (moderate concern, but not classic for malignancy). To determine the clinical significance of BI-RADS 4 subcategories and the common pathologic changes associated with these mammographic lesions, a retrospective review of 239 consecutive stereotactic-needle core biopsies (SNCB) for microcalcifications was performed. All 239 SNCBs were BI-RADS 4 lesions, and of these, 191 were subcategorized to 4A, 4B or 4C. Ninety-four of 191 (49%) were 4A, 73 (38%) were 4B, and 24 (13%) were 4C. Fibrocystic change was the most common finding (66/239; 28%) followed by ductal carcinoma in situ (DCIS) accounting for 23% of cases. This was followed by columnar cell alteration with or without atypia (47/239; 19%), and fibroadenoma (45/239; 19%). While 70% (17/24) of BI-RADS 4C category lesions were DCIS, only 21% (15/73) of BI-RADS 4B and 10% (10/94) of BI-RADS 4A were DCIS. Without sub-categorization, carcinoma was diagnosed in 23% (55/239) of all cases with BI-RADS 4. Therefore, subcategorizing BI-RADS 4 lesions is important since it not only benefits the patient and clinician in understanding the level of concern for carcinoma, but will also alert the pathologist.

A unique case of cystic hypersecretory carcinoma in a 48-year-old woman who presented with a tender, palpable abnormality of the lower outer quadrant of her left breast in 2002 is described. Mammographic and ultrasound examinations showed heterogeneous, dense breast tissue with a focal asymmetric density and an ill-defined heterogenous hypoechoic area with anechoic spaces, respectively, that corresponded to the area of palpable abnormality. Although the imaging findings did not change significantly over 4 years, the patient complained of intermittent, spontaneous serous discharge from the left nipple and persistent induration and tenderness in the same area of the left breast. An ultrasound-guided core-needle biopsy of the lesion was performed at the last visit to rule out a malignant process. A left simple mastectomy showed a large multicystic lesion occupying most of the lower quadrants by a cystic hypersecretory carcinoma and Paget disease of the nipple.

The phosphatidylinositol 3-kinase (PI3-K) signalling pathway has been implicated in breast cancer development and resistance to therapy. Akt-1 and serum and glucocorticoid-induced kinase-1 (SGK-1) are homologous kinases which are important downstream effectors of PI3-K signalling. We sought to determine the individual expression patterns of these two kinases in order to better understand their respective roles in PI3-K signalling in breast cancer. To this end, we examined the expression of both p-Akt-1 and SGK-1 in 40 breast cancers. p-Akt-1 expression was seen in 58% of tumour samples, while SGK-1 overexpression was detected in 48%. Interestingly, a highly significant association was found between the expression of p-Akt-1 and SGK-1 (P=0.002), suggesting complementary physiological functions in PI3-K signalling. This finding is consistent with recent genetic data from Caenorhabditis elegans suggesting that both SGK-1 and Akt-1 are required for signalling downstream of insulin receptor activation.

Background: This study sought to determine whether the type of biopsy examination independently affects sentinel lymph node (SLN) status in breast cancer patients. Methods: A prospective multicenter study of patients who had SLN biopsy examination followed by axillary node dissection was analyzed to determine whether the type of biopsy examination influenced SLN status. Results: Of the 3853 patients studied, 32% had a positive SLN. Patients were diagnosed by fine-needle (N = 293), core-needle (N = 2154), excisional (N = 1386), or incisional (N = 20) biopsy procedures. The rates of SLN positivity for these groups were 45%, 32%, 29%, and 65%, respectively (P

α-Methylacyl-coenzyme A racemase (AMACR) is a sensitive and specific tissue marker for the diagnosis of prostatic carcinoma. However, limited data are available on AMACR expression in residual prostatic carcinoma following hormone therapy. We analyzed 64 residual or recurrent prostatic adenocarcinomas following hormonal therapy for the expression of AMACR using a monoclonal antibody (P504S) to AMACR. In 20 localized cases, AMACR staining was absent in 11 (55%), 1+ in 6 (30%), and 2+ or 3+ in 3 (15%). However, in 15 metastatic cases, AMACR was absent in 1 (7%), 1+ in 3 (20%), and 2+ or 3+ in 11 (73%). None of the 29 postradiotherapy cases showed complete absence of AMACR staining: 2 (7%) were 1+, and 27 (93%) were 2+ or 3+. AMACR expression was reduced significantly in the majority of posthormonal residual carcinomas, whereas in postradiotherapy and in hormone-refractory metastatic prostatic adenocarcinoma, AMACR expression was retained. Therefore, the diagnosis of residual prostatic carcinoma after hormonal therapy using AMACR immunostaining must be interpreted with caution. Furthermore, AMACR might have a role in the recurrence of prostatic adenocarcinoma after medical therapy.

Atypical cardiac myxomas are a rare occurrence and may present with a variety of clinical manifestations depending on the morphology and location. We present three cases of myxomas located atypically on the left atrial appendage ridge, chordae of the mitral valve, and right atrium. Echocardiography plays a prominent role in the diagnosis and follow-up of these patients. (ECHOCARDIOGRAPHY, Volume 21, January 2004)

Objective To evaluate several conventional cervical smears obtained from women undergoing routine screening for cervical dysplasia or carcinoma and whose smears contained structures resembling collagen balls. Study design Between 1995 and 1998, cervical smears containing collagen balls were analyzed. The clinical histories of the patients whose smears contained collagen balls, including the gestational history, were reviewed. Histopathologic material from any related surgical specimens was reviewed, with special attention to mesothelial surfaces. Results Collagen balls were found in 5 of 77,891 Pap smears examined (0.006%). None of the patients had evidence of neoplasms of the genital tract. One of the patients was in the first trimester of pregnancy. Conclusion We suggest that collagen balls in cervical smear originate in mesothelium-covered organs, from where they are transported via the fallopian tubes into the uterine cavity. Their significance lies in their being mistaken for mucin-distended cells exfoliated from a neoplasm or from detached fragments of a papillary ovarian neoplasm.

Anaplastic large cell lymphoma is a rare type of primary breast lymphoma. We report a case of anaplastic large cell lymphoma, T-cell phenotype, occurring in the periprosthetic capsule of a silicone breast prosthesis 9 years after implantation for augmentation mammoplasty. This case is unique for its unusual presentation.

Various nonneoplastic processes and benign tumors can simulate lung cancer at different levels of diagnostic work-up: during physical examination, interpretation of radiologic images, or examination of pathologic material. Despite the advances in imaging studies, radiologic findings at times can be misleading due to an overlap in the appearance between neoplastic and nonneoplastic lesions. Cytologic evaluation and small tissue biopsy have become established initial methods for the evaluation of radiographically detected lung lesions. These methods are less invasive compared to video-assisted thoracic surgery, thoracotomy, and mediastinoscopy. However, specimens obtained from these methods are smaller than those obtained from the invasive procedures and can occasionally pose diagnostic challenges potentially resulting in misdiagnosis. Therefore, familiarity with the potential mimickers of lung carcinoma is of utmost importance.

To evaluate the expression of cytokeratin (CK) 19, we stained sections obtained from formalin-fixed, paraffin tissue blocks of 35 thyroid tumors (follicular adenoma [FA], 20; papillary thyroid carcinoma [PTC], 10 follicular variant [FV] and 5 usual type) and scored the extent of staining as follows: 1+ (5% positively stained cells), 2+ (5%-25% positively stained cells), 3+ (25%-75% positively stained cells), and 4+ (75% positively stained cells). All 15 PTCs (including 10 FV-PTCs) were CK19 positive: 14 were 4+ and 1 (FV-PTC) was 2+. All 20 FAs also were CK19 positive: 15 were 1+, 1 was 2+, 4 were 3+, and none was 4+. In the FAs that were scored 1+, reactivity usually was confined to follicular cells lining cystically dilated atrophic follicles that lacked the typical nuclear features of PTC. The remaining FAs showed more diffuse reactivity, which was, however, less intense than that observed in the PTCs. Thus, immunoreactivity for CK19 is not specific for PTC, although we acknowledge that the extent and intensity of staining are considerably greater in this tumor than in FA. There were no significant differences in staining for CK19 between nonneoplastic follicles adjacent to PTCs and those adjacent to FAs.