Identification of Glandular (Acinar)/Tubule Formation in Invasive Carcinoma of the Breast: A Study to Determine Concordance Using the World Health Organization Definition.

CONTEXT.­: The Nottingham Grading System (NGS) developed by Elston and Ellis is used to grade invasive breast cancer (IBC). Glandular (acinar)/tubule formation is a component of NGS. OBJECTIVE.­: To investigate the ability of pathologists to identify individual structures that should be classified as glandular (acinar)/tubule formation. DESIGN.­: A total of 58 hematoxylin-eosin photographic images of IBC with 1 structure circled were classified as tubules (41 cases) or nontubules (17 cases) by Professor Ellis. Images were sent as a PowerPoint (Microsoft) file to breast pathologists, who were provided with the World Health Organization definition of a tubule and asked to determine if a circled structure represented a tubule. RESULTS.­: Among 35 pathologists, the κ statistic for assessing agreement in evaluating the 58 images was 0.324 (95% CI, 0.314-0.335). The median concordance rate between a participating pathologist and Professor Ellis was 94.1% for evaluating 17 nontubule cases and 53.7% for 41 tubule cases. A total of 41% of the tubule cases were classified correctly by less than 50% of pathologists. Structures classified as tubules by Professor Ellis but often not recognized as tubules by pathologists included glands with complex architecture, mucinous carcinoma, and the "inverted tubule" pattern of micropapillary carcinoma. A total of 80% of participants reported that they did not have clarity on what represented a tubule. CONCLUSIONS.­: We identified structures that should be included as tubules but that were not readily identified by pathologists. Greater concordance for identification of tubules might be obtained by providing more detailed images and descriptions of the types of structures included as tubules.

Emerging Landscape of Targeted Therapy of Breast Cancers With Low Human Epidermal Growth Factor Receptor 2 Protein Expression.

CONTEXT.­: Human epidermal growth factor receptor 2 (HER2) status in breast cancer is currently classified as negative or positive for selecting patients for anti-HER2 targeted therapy. The evolution of the HER2 status has included a new HER2-low category defined as an HER2 immunohistochemistry score of 1+ or 2+ without gene amplification. This new category opens the door to a targetable HER2-low breast cancer population for which new treatments may be effective. OBJECTIVE.­: To review the current literature on the emerging category of breast cancers with low HER2 protein expression, including the clinical, histopathologic, and molecular features, and outline the clinical trials and best practice recommendations for identifying HER2-low-expressing breast cancers by immunohistochemistry. DATA SOURCES.­: We conducted a literature review based on peer-reviewed original articles, review articles, regulatory communications, ongoing and past clinical trials identified through ClinicalTrials.gov, and the authors' practice experience. CONCLUSIONS.­: The availability of new targeted therapy potentially effective for patients with breast cancers with low HER2 protein expression requires multidisciplinary recognition. In particular, pathologists need to recognize and identify this category to allow the optimal selection of patients for targeted therapy.

Analytical Validation of the PreciseDx Digital Prognostic Breast Cancer Test in Early-Stage Breast Cancer.

BACKGROUND: PreciseDx Breast (PDxBr) is a digital test that predicts early-stage breast cancer recurrence within 6-years of diagnosis. MATERIALS AND METHODS: Using hematoxylin and eosin-stained whole slide images of invasive breast cancer (IBC) and artificial intelligence-enabled morphology feature array, microanatomic features are generated. Morphometric attributes in combination with patient's age, tumor size, stage, and lymph node status predict disease free survival using a proprietary algorithm. Here, analytical validation of the automated annotation process and extracted histologic digital features of the PDxBr test, including impact of methodologic variability on the composite risk score is presented. Studies of precision, repeatability, reproducibility and interference were performed on morphology feature array-derived features. The final risk score was assessed over 20-days with 2-operators, 2-runs/day, and 2-replicates across 8-patients, allowing for calculation of within-run repeatability, between-run and within-laboratory reproducibility. RESULTS: Analytical validation of features derived from whole slide images demonstrated a high degree of precision for tumor segmentation (0.98, 0.98), lymphocyte detection (0.91, 0.93), and mitotic figures (0.85, 0.84). Correlation of variation of the assay risk score for both reproducibility and repeatability were less than 2%, and interference from variation in hematoxylin and eosin staining or tumor thickness was not observed demonstrating assay robustness across standard histopathology preparations. CONCLUSION: In summary, the analytical validation of the digital IBC risk assessment test demonstrated a strong performance across all features in the model and complimented the clinical validation of the assay previously shown to accurately predict recurrence within 6-years in early-stage invasive breast cancer patients.

Correlation of the Ki67 Working Group prognostic risk categories with the Oncotype DX Recurrence Score in early breast cancer.

BACKGROUND: The relationship between Ki67 assessed by immunohistochemistry (IHC) and the Oncotype DX Recurrence Score (RS) is unclear. The objective of this study was to determine the correlation between the 21-gene RS and IHC-measured Ki67 with the prognostic classification groups recommended by the International Ki67 Working Group (IKWG). METHODS: The authors performed a retrospective chart review of women who had hormone receptor (HR)-positive, human epidermal growth factor receptor 2-negative early breast cancer with zero to three positive lymph nodes and both Ki67 and the 21-gene RS performed at their institution from 2013 to 2021. Patients were categorized into low (≤5%), intermediate (6%-29%), and high Ki67 groups (≥30%) according to IKWG recommendations. Overall agreement and risk-stratified agreement between Ki67 and RS were assessed with the proportion of agreement and the κ statistic. RESULTS: The study included 525 patients with HR-positive breast cancer. Among the 49% of patients with intermediate Ki67 values of 6%-29%, the distribution of low (0-10), intermediate (11-25), and high RS (26-100) was 19%, 66%, and 15%, respectively. There was slight agreement (κ = 0.01-0.20) between Ki67 and RS (κ = 0.027) in the overall population, although this was not significant (p = .1985). There was fair agreement (κ = 0.21-0.40) between high Ki67 and RS values (κ = 0.280; p < .0001). A higher progesterone receptor percentage was associated with lower RS values (p > .0001) but not lower Ki67 values. A positive nodal status and a larger tumor size were associated with higher Ki67 values (p = .0059 and p < .0001) but not with RS. CONCLUSIONS: In this group of patients selected to have a 21-gene RS, there was no significant correlation between Ki67 and RS in the overall population, and there was fair agreement between high Ki67 and high RS values. LAY SUMMARY: In patients with early-stage, hormone receptor-positive breast cancer, decisions on adjuvant chemotherapy are based on certain biological features of the cancer and genomic assays such as the Oncotype DX Recurrence Score (RS). The goal of this study was to determine the correlation between Ki67, a marker of proliferation, and the Oncotype DX RS, a 21-gene assay demonstrated to be predictive of an adjuvant chemotherapy benefit in patients with early-stage breast cancer. In 525 patients, the authors did not find a significant correlation between Ki67 and RS.

Concordance in Breast Cancer Grading by Artificial Intelligence on Whole Slide Images Compares With a Multi-Institutional Cohort of Breast Pathologists.

CONTEXT.­: Breast carcinoma grade, as determined by the Nottingham Grading System (NGS), is an important criterion for determining prognosis. The NGS is based on 3 parameters: tubule formation (TF), nuclear pleomorphism (NP), and mitotic count (MC). The advent of digital pathology and artificial intelligence (AI) have increased interest in virtual microscopy using digital whole slide imaging (WSI) more broadly. OBJECTIVE.­: To compare concordance in breast carcinoma grading between AI and a multi-institutional group of breast pathologists using digital WSI. DESIGN.­: We have developed an automated NGS framework using deep learning. Six pathologists and AI independently reviewed a digitally scanned slide from 137 invasive carcinomas and assigned a grade based on scoring of the TF, NP, and MC. RESULTS.­: Interobserver agreement for the pathologists and AI for overall grade was moderate (κ = 0.471). Agreement was good (κ = 0.681), moderate (κ = 0.442), and fair (κ = 0.368) for grades 1, 3, and 2, respectively. Observer pair concordance for AI and individual pathologists ranged from fair to good (κ = 0.313-0.606). Perfect agreement was observed in 25 cases (27.4%). Interobserver agreement for the individual components was best for TF (κ = 0.471 each) followed by NP (κ = 0.342) and was worst for MC (κ = 0.233). There were no observed differences in concordance amongst pathologists alone versus pathologists + AI. CONCLUSIONS.­: Ours is the first study comparing concordance in breast carcinoma grading between a multi-institutional group of pathologists using virtual microscopy to a newly developed WSI AI methodology. Using explainable methods, AI demonstrated similar concordance to pathologists alone.

Mastectomy or Margin Re-excision? A Nomogram for Close/Positive Margins After Lumpectomy for DCIS.

BACKGROUND: Anatomic extent of ductal carcinoma in situ (DCIS) may be uncertain in spite of clinical, pathologic, and imaging data. Consequently close/positive margins are common with lumpectomy for DCIS and often lead to a challenge in deciding whether to perform a re-excision or mastectomy. PATIENTS AND METHODS: From a single health system, we identified cases of lumpectomy for DCIS with close/positive margins who underwent re-excision for the purpose of constructing a nomogram. In total, 289 patients were available for analysis. The patients were randomly divided into two sets allocating 70% to the modeling and 30% to the validation set. A multivariable logistic regression model was used to estimate the probability of overall positive margin status using multiple clinicopathologic predictors. Nomogram validation included internal tenfold cross-validation, internal bootstrap validation, and external validation for which a concordance index was calculated to assess the external validity. RESULTS: Significant predictors of persistent positive margins from regression modeling included necrosis at diagnosis (non-comedo or comedo); DCIS not associated with calcifications on core biopsy; high-grade DCIS; progesterone receptor positivity; and number of positive margins at initial surgery. When subjected to internal validation, the nomogram achieved an uncorrected concordance index of 0.7332, a tenfold cross-validation concordance index of 0.6795, and a bootstrap-corrected concordance index of 0.6881. External validation yielded an estimated concordance index of 0.7095. CONCLUSION: Using clinical and pathologic variables from initial diagnosis and surgery for DCIS, this nomogram predicts persistent positive margins with margin re-excision, and may be a valuable tool in surgical decision-making.

Does previous history of cancer or atypia predict histologic upgrade for pure intraductal papillomas diagnosed via core biopsy? A study of 490 cases at a single institution.

BACKGROUND: Management of pure intraductal papillomas (IDP) without atypia diagnosed on core needle biopsy (CNB) remains controversial given highly variable rates of upgrade in the literature. AIM: We sought to identify clinical and histologic factors that predict upgrade to atypia or malignancy in a large population. METHODS AND RESULTS: A retrospective review was performed of all cases of pure IDP diagnosed on CNB and then surgically excised at a single institution from 2008 to 2018. Clinical, radiologic, and pathologic factors were compared in the no upgrade, upgrade to atypia, or upgrade to cancer groups. Univariate analysis was performed comparing no upgrade and upgrade to cancer or atypia. Four hundred and thirty nine patients were identified with a total of 490 IDP and a median age of 50 years (range 16-85). Of these patients, 54 (12.3%) were upgraded to atypia after surgical excision and five (1.1%) were upgraded to cancer. The presence of multiple papillomas in a single patient was a significant predictor of upgrade to cancer or atypia (p < .01), as well as age over ≥55 years (p < .01) and a prior history of cancer (p < .01). No other clinical, radiologic and histologic factors were found to be significant predictors of upgrade. 40/439 (9.1%) patients in the total cohort had prior history of cancer, and of these, 2/40 (5%) were found to have a new cancer after excision. CONCLUSIONS: In patients with pure IDP on CNB, the upgrade rate to malignancy was 1.1%, while 12.3% were upgraded to atypia. The clinical significance of identifying atypia in a papilloma is unknown, especially in a patient with a prior history of atypia or cancer. However, the majority of patients who were upgraded to either atypia or cancer had no prior history of high-risk or malignant breast disease and are therefore considered true clinical upgrades. As such excision for IDP should be considered.

High endogenous CCL2 expression promotes the aggressive phenotype of human inflammatory breast cancer.

Inflammatory Breast Cancer (IBC) is a highly aggressive malignancy with distinct clinical and histopathological features whose molecular basis is unresolved. Here we describe a human IBC cell line, A3250, that recapitulates key IBC features in a mouse xenograft model, including skin erythema, diffuse tumor growth, dermal lymphatic invasion, and extensive metastases. A3250 cells express very high levels of the CCL2 chemokine and induce tumors enriched in macrophages. CCL2 knockdown leads to a striking reduction in macrophage densities, tumor proliferation, skin erythema, and metastasis. These results establish IBC-derived CCL2 as a key factor driving macrophage expansion, and indirectly tumor growth, with transcriptomic analysis demonstrating the activation of multiple inflammatory pathways. Finally, primary human IBCs exhibit macrophage infiltration and an enriched macrophage RNA signature. Thus, this human IBC model provides insight into the distinctive biology of IBC, and highlights potential therapeutic approaches to this deadly disease.

Secondary Invasive Breast Events among Patients with Hormone-Positive Breast Cancer and High-Risk Oncotype DX Recurrence Scores 26-30 and ≥31.

BACKGROUND: The Oncotype DX Recurrence Score (ODx RS) is the most widely adopted genomic assay used to guide treatment for patients with early-stage, hormone-positive (HR+) breast cancer (BC), with higher scores predicting greater risk of recurrence and benefit from chemotherapy. Patients with ODx RS >25 typically recieve adjuvant chemotherapy; however, data regarding efficacy of chemotherapy for reducing recurrence in this population have been mixed. OBJECTIVES: This study aimed to evaluate outcomes of patients with early-stage HR+ BC with high-risk ODx RS (26-30 and ≥31) in order to assess treatment patterns and outcomes. We hypothesized that the benefit of chemotherapy in these groups may be minimal and that select patients may forgo chemotherapy in favor of more aggressive endocrine therapy and ovarian suppression. METHODS: We performed a retrospective analysis of 515 patients with early-stage, HR+ BC with high-risk ODx RS 26-30 and ≥31 treated between 2006 and 2018. Patients were stratified by RS: low-risk (≤10), intermediate-risk (11-25), and high-risk (≥26). The Kaplan-Meier method was used to estimate the time to secondary invasive breast events (SIBE) or distributions overall and among different RS groups with the log rank test used to compare distributions between groups. RESULTS: Rates of chemotherapy administration were 7% among the low-risk group, 18% among the intermediate-risk group, and 83% among high-risk patients with 41 SIBE (8%) reported. When stratified by ODx RS, 5-year rates of SIBE were 4%, 6%, and 16% for low-risk, intermediate-risk, and high-risk RS, respectively. Among the 27 lymph node (LN)-negative patients with ODx RS 26-30, 74% received chemotherapy. The 5-year rate of SIBE was 25% among patients who received chemotherapy and 33% among those who did not (p = 0.5489). Among the 23 LN-negative patients with ODx RS ≥31, 91% of patients received chemotherapy. The 5-year rate of SIBE was 0% both with and without chemotherapy. CONCLUSIONS: There was no statistically significant difference in SIBE for patients with high-risk ODx RS based on chemotherapy treatment. More aggressive endocrine therapy with ovarian suppression has become an alternative to chemotherapy among patients with intermediate-risk ODx RS (16-25). This approach may be useful among patients with high-risk ODx RS, with additional studies needed in this patient population.

Interobserver variability in the assessment of stromal tumor-infiltrating lymphocytes (sTILs) in triple-negative invasive breast carcinoma influences the association with pathological complete response: the IVITA study.

High stromal tumor-infiltrating lymphocytes (sTILs) in triple-negative breast cancer (TNBC) are associated with pathological complete response (pCR) after neoadjuvant chemotherapy (NAC). Histopathological assessment of sTILs in TNBC biopsies is characterized by substantial interobserver variability, but it is unknown whether this affects its association with pCR. Here, we aimed to investigate the degree of interobserver variability in an international study, and its impact on the relationship between sTILs and pCR. Forty pathologists assessed sTILs as a percentage in digitalized biopsy slides, originating from 41 TNBC patients who were treated with NAC followed by surgery. Pathological response was quantified by the MD Anderson Residual Cancer Burden (RCB) score. Intraclass correlation coefficients (ICCs) were calculated per pathologist duo and Bland-Altman plots were constructed. The relation between sTILs and pCR or RCB class was investigated. The ICCs ranged from -0.376 to 0.947 (mean: 0.659), indicating substantial interobserver variability. Nevertheless, high sTILs scores were significantly associated with pCR for 36 participants (90%), and with RCB class for eight participants (20%). Post hoc sTILs cutoffs at 20% and 40% resulted in variable associations with pCR. The sTILs in TNBC with RCB-II and RCB-III were intermediate to those of RCB-0 and RCB-I, with lowest sTILs observed in RCB-I. However, the limited number of RCB-I cases precludes any definite conclusions due to lack of power, and this observation therefore requires further investigation. In conclusion, sTILs are a robust marker for pCR at the group level. However, if sTILs are to be used to guide the NAC scheme for individual patients, the observed interobserver variability might substantially affect the chance of obtaining a pCR. Future studies should determine the 'ideal' sTILs threshold, and attempt to fine-tune the patient selection for sTILs-based de-escalation of NAC regimens. At present, there is insufficient evidence for robust and reproducible sTILs-guided therapeutic decisions.

Response in breast vs axilla after neoadjuvant treatment and implications for nonoperative management of invasive breast cancer.

Improved imaging and neoadjuvant chemotherapy (NAT) have led to higher pathologic complete response rates (pCR) in patients with invasive breast cancer. This has questioned the necessity of surgery and axillary lymph node (ALN) dissection in these patients. Prospective clinical trials are implementing extensive core biopsies of the tumor bed of patients with clinical complete response as a means to identify and spare them breast surgery. In addition, it is anticipated that patients with pCR are most likely going to have no or minimal disease in ALN as well. To verify the feasibility of these trials, we performed a pathologic analysis of all our patients who have undergone NAT from 2009 to present. Using pathology data base, we identified 362 patients treated with neoadjuvant chemotherapy followed by surgery. Clinical and pathologic information including gross and microscopic descriptions as well as biomarker status was collected. pCR was 50% for patients with negative ALN pretreatment but only 28% for patients with positive ALN at diagnosis. Despite achieving pCR in the breast, up to 10% of patients with positive ALN and 1% with negative ALN had persistent disease. Eight percent of patients that were presumed to have no ALN disease either clinically and or by imaging were found to have metastatic carcinoma in ALN. The metastases were predominantly (80%) <5 mm, and not palpable on physical examination and or due to biopsy sampling error. pCR in breast and ALN directly correlated with tumor size, ALN disease, and Her2 positive and triple negative receptor phenotype. In breast cancer patients who are node positive at time of diagnosis with pCR in the breast after neoadjuvant chemotherapy, residual lymph node disease was very uncommon. Further study is warranted to select patients who may avoid breast and axillary surgery post neoadjuvant chemotherapy.

Measuring Invasive Breast Carcinoma on Core Biopsy-Is It Necessary?

CONTEXT.­: Pathologic tumor size is significant in the treatment of breast carcinoma and is routinely measured on excision. OBJECTIVE.­: To analyze the need for measuring size of invasive mammary carcinoma on biopsy. DESIGN.­: Nine hundred twenty-two cases of invasive carcinoma whose size was measured (greatest linear measurement) on biopsy and excision was correlated, including imaging when available (110 cases). RESULTS.­: Patient mean age was 62 years. Most (90%; 830 of 922) carcinomas were ductal and sampled by ultrasound and graded as follows: well, 13% (113 of 922); moderately, 58% (532 of 922), and poorly differentiated, 28% (258 of 922); 19 microinvasive not graded. Tumor mean size was 7.5 mm on biopsy and 14.4 mm on excision. Biopsy modality was as follows: ultrasound, 7.8 mm (92%, 844 of 922); mammotome, 3.3 mm (7%, 65 of 922); and magnetic resonance imaging, 5.9 mm (1%, 13 of 922). Size comparison on biopsy versus excision was biopsy > excision: 8% (72 of 922), biopsy = excision: 10% (95 of 922), and biopsy < excision: 82% (755 of 922). Half (36 of 72) of the biopsy > excision tumors were less than 5 mm, 96% (726 of 755) of biopsy < excision tumors were greater than 5 mm, while those equal on both were predominantly (88%, 84 of 95) less than 10 mm, 20% (19 of 95) of which were microinvasive. Stage changed in 600 cases, staging based on excision in 581 (63%), and staging based on biopsy in 19 (2%). Radiologic-pathologic correlation (n = 110) showed perfect concordance in 11 (10%), 83 (75%) were ±1 to 2 mm, and 16 (15%) were ± more than 3 mm. Difference between the biopsy and excision ranged from a lower limit of 1.3 mm for T1a tumors to 18 mm for T2. CONCLUSIONS.­: While most carcinomas are larger on excision, 18% (167 of 922) are larger or equal on biopsy. Factors predictive of biopsy > excision tumors include stage 1 tumors (P < .001), especially less than 5 mm, and sampled by mammotome. We recommend measuring invasive carcinoma on biopsy and excision.

Histologic grading of breast carcinoma: a multi-institution study of interobserver variation using virtual microscopy.

Breast carcinoma grading is an important prognostic feature recently incorporated into the AJCC Cancer Staging Manual. There is increased interest in applying virtual microscopy (VM) using digital whole slide imaging (WSI) more broadly. Little is known regarding concordance in grading using VM and how such variability might affect AJCC prognostic staging (PS). We evaluated interobserver variability amongst a multi-institutional group of breast pathologists using digital WSI and how discrepancies in grading would affect PS. A digitally scanned slide from 143 invasive carcinomas was independently reviewed by 6 pathologists and assigned grades based on established criteria for tubule formation (TF), nuclear pleomorphism (NP), and mitotic count (MC). Statistical analysis was performed. Interobserver agreement for grade was moderate (κ = 0.497). Agreement was fair (κ = 0.375), moderate (κ = 0.491), and good (κ = 0.705) for grades 2, 3, and 1, respectively. Observer pair concordance ranged from fair to good (κ = 0.354-0.684) Perfect agreement was observed in 43 cases (30%). Interobserver agreement for the individual components was best for TF (κ = 0.503) and worst for MC (κ = 0.281). Seventeen of 86 (19.8%) discrepant cases would have resulted in changes in PS and discrepancies most frequently resulted in a PS change from IA to IB (n = 9). For two of these nine cases, Oncotype DX results would have led to a PS of 1A regardless of grade. Using VM, a multi-institutional cohort of pathologists showed moderate concordance for breast cancer grading, similar to studies using light microscopy. Agreement was the best at the extremes of grade and for evaluation of TF. Whether the higher variability noted for MC is a consequence of VM grading warrants further investigation. Discordance in grading infrequently leads to clinically meaningful changes in the prognostic stage.

MRI Features of Free Liquid Silicone in the Transgender Female Breast.

OBJECTIVE: To define MRI features of free liquid silicone injection (FLSI) of the breast in transgender women considering surgical management. METHODS: This study was IRB-approved. MRI images from transgender women with FLSI imaged between 2009 and 2019 were reviewed. Presence and location of fibrotic masses (FMs) in the breast(s) and pectoralis muscle and patterns of granulomas were correlated with clinicopathologic findings. Background enhancement was quantified. Comparisons were performed using two-tailed Fisher exact and Student's t test. RESULTS: Of 21 transgender women with FLSI (mean age 46.8 years), 13/21 (61.9%) had a dominant FM measuring over 4 cm; these were limited to breast and pectoralis in 6/21 (28.6%), breast in 9/21 (42.9%), and pectoralis only in 2/21 (9.5%). Four of 21 patients (19.0%) had no FMs, and 4/21 (19.0%) had masses under 4 cm. Mean size of the dominant FM was 7.4 cm (range 4-12 cm). FMs were enhancing in 5/13 (38.5%) and contained T2 high signal granulomas in 8/13 (61.5%). While 18/21 (85.8%) of cases showed mild to moderate overall background enhancement, the majority 7/13 (61.5%) of dominant FM were non-enhancing. About half of cases (11/21, 52.4%) had diffuse foci, and half (10/21, 47.6%) had diffuse foci and masses throughout the breast and pectoralis muscle. These foci and masses displayed T2 high signal in 13/21 (61.9%). There were no occult carcinomas observed. CONCLUSION: MRI performed on symptomatic FLSI patients considering surgical treatment is helpful in assessing the extent of silicone infiltration and fibrotic reaction of the breast and pectoralis muscle.

Interobserver variability in upfront dichotomous histopathological assessment of ductal carcinoma in situ of the breast: the DCISion study.

Histopathological assessment of ductal carcinoma in situ, a nonobligate precursor of invasive breast cancer, is characterized by considerable interobserver variability. Previously, post hoc dichotomization of multicategorical variables was used to determine the "ideal" cutoffs for dichotomous assessment. The present international multicenter study evaluated interobserver variability among 39 pathologists who performed upfront dichotomous evaluation of 149 consecutive ductal carcinomas in situ. All pathologists independently assessed nuclear atypia, necrosis, solid ductal carcinoma in situ architecture, calcifications, stromal architecture, and lobular cancerization in one digital slide per lesion. Stromal inflammation was assessed semiquantitatively. Tumor-infiltrating lymphocytes were quantified as percentages and dichotomously assessed with a cutoff at 50%. Krippendorff's alpha (KA), Cohen's kappa and intraclass correlation coefficient were calculated for the appropriate variables. Lobular cancerization (KA = 0.396), nuclear atypia (KA = 0.422), and stromal architecture (KA = 0.450) showed the highest interobserver variability. Stromal inflammation (KA = 0.564), dichotomously assessed tumor-infiltrating lymphocytes (KA = 0.520), and comedonecrosis (KA = 0.539) showed slightly lower interobserver disagreement. Solid ductal carcinoma in situ architecture (KA = 0.602) and calcifications (KA = 0.676) presented with the lowest interobserver variability. Semiquantitative assessment of stromal inflammation resulted in a slightly higher interobserver concordance than upfront dichotomous tumor-infiltrating lymphocytes assessment (KA = 0.564 versus KA = 0.520). High stromal inflammation corresponded best with dichotomously assessed tumor-infiltrating lymphocytes when the cutoff was set at 10% (kappa = 0.881). Nevertheless, a post hoc tumor-infiltrating lymphocytes cutoff set at 20% resulted in the highest interobserver agreement (KA = 0.669). Despite upfront dichotomous evaluation, the interobserver variability remains considerable and is at most acceptable, although it varies among the different histopathological features. Future studies should investigate its impact on ductal carcinoma in situ prognostication. Forthcoming machine learning algorithms may be useful to tackle this substantial diagnostic challenge.

Evaluation of surgically excised breast tissue microstructure using wide-field optical coherence tomography.

BACKGROUND: Currently, positive margins at lumpectomy contribute to health care cost, patient anxiety, and treatment delay. Multiple technology solutions are being explored with the aim of lowering re-excision rates for breast-conserving surgery (BCS). We examined wide-field optical coherence tomography (WF-OCT), an innovative adjunct intraoperative imaging tool for tissue visualization of margins. METHODS: This IRB-approved pilot study included women with invasive or in situ carcinoma scheduled for primary BCS. Lumpectomy specimens and any final/revised margins were imaged by optical coherence tomography immediately prior to standard histological processing. The optical coherence tomography used provided two-dimensional, cross-sectional, real-time depth visualization of the margin widths around excised specimens. A volume of images was captured for 10 × 10 cm tissue surface at high resolution (sub-30 µm) to a depth of 2 mm. Integrated interpretation was performed incorporating final pathology linked with the optical image data for correlation. RESULTS: Wide-field optical coherence tomography was performed on 185 tissue samples (50 lumpectomy specimens and 135 additional margin shaves) in 50 subjects. Initial diagnosis was invasive ductal carcinoma (IDC) in 10, ductal carcinoma in situ (DCIS) in 14, IDC/DCIS in 22, invasive lobular carcinoma (ILC) in 2, ILC/DCIS in 1, and sarcoma in 1. Optical coherence tomography was concordant with final pathology in 178/185 tissue samples for overall accuracy of 86% and 96.2% (main specimen alone and main specimen + shave margins). Of seven samples that were discordant, 57% (4/7) were considered close (DCIS < 2 mm from margin) per final pathology. CONCLUSION: Wide-field optical coherence tomography demonstrated concordance with histology at tissue margins, supporting its potential for use as a real-time adjunct intraoperative imaging tool for margin assessment. Further studies are needed for comprehensive evaluation in the intraoperative setting.

Adenomyoepitheliomas of the Breast Frequently Harbor Recurrent Hotspot Mutations in PIK3-AKT Pathway-related Genes and a Subset Show Genetic Similarity to Salivary Gland Epithelial-Myoepithelial Carcinoma.

Adenomyoepitheliomas (AME) of the breast and epithelial-myoepithelial carcinomas (EMCs) of salivary gland are morphologically similar tumors defined by the presence of a biphasic population of ductal epithelial elements mixed with myoepithelial cells. We sought to explore the molecular profile of AMEs and determine whether they might also share the PLAG1, HMGA2, and HRAS alterations seen in EMCs. Tumor tissue from 19 AMEs was sequenced and analyzed using Ion AmpliSeq Cancer Hotspot Panel v2 covering ∼2800 COSMIC mutations across 50 cancer-related genes. Cases were additionally screened by FISH for PLAG1 and HMGA2 rearrangements. Of 19 AMEs (12 benign; 7 malignant), 2 cases failed the DNA extraction. Of the remaining 17 cases, 14 had at least one nonsynonymous mutation identified. The most common mutations were in PIK3CA (6/17) and AKT1 (5/17), which were mutually exclusive. Two tumors demonstrated mutations in APC, while 1 demonstrated an STK11 mutation. Mutations in ATM, EGFR, FGFR3 or GNAS were identified in 4 cases with concurrent AKT1 mutations. HRAS mutation co-occurring with PIK3CA mutation was noted in 1 case of ER-negative malignant AME. While 2 cases harbored alterations in HMGA2, none was positive for PLAG1 rearrangement. Our findings confirm that breast AMEs are genetically heterogeneous exhibiting recurrent mutually exclusive mutations of PIK3CA and AKT1 in a majority of cases. HRAS mutations co-occur with PIK3CA mutations in ER-negative AMEs and may possibly be linked to clinically aggressive behavior. We identified hotspot mutations in additional genes (APC, STK11, ATM, EGFR, FGFR3, and GNAS). We report the presence of HMGA2 alterations in 2/16 AMEs, supporting their relationship with EMC of salivary glands in at least a subset of cases. PIK3CA, AKT1 and HRAS may serve as potential actionable therapeutic targets in clinically aggressive AMEs.

Not All Ductal Carcinomas In Situ Are Created IDLE (Indolent Lesions of Epithelial Origin).

CONTEXT.­: Mammographic screening has increased the incidence of ductal carcinoma in situ (DCIS), but this has not been accompanied by a decline in the incidence of invasive carcinoma (IC). Consequently, the surgical treatment of DCIS has recently been questioned, with some advocating only surveillance (with or without neoadjuvant endocrine therapy) after a core biopsy diagnosis of DCIS. OBJECTIVES.­: To examine the predictive value of a core biopsy diagnosis of DCIS, particularly the upgrade rate to IC, and to identify associated factors. DESIGN.­: Using the pathology database, we identified 2943 cases of DCIS diagnosed on core biopsy from 2000 to 2015, of which 229 cases (8%) later had the stage upgraded to IC. RESULTS.­: Ages ranged from 25 to 90 years (mean, 59 years). The DCIS presented with calcifications in 151 of 229 cases (65.9%), was widespread in 26 of 151 cases (17%), had a mass or density present in 70 of 229 cases (31%), with heterogeneous echogenic features in 44 of those 70 cases (63%), and an enhancement upon magnetic resonance imaging in 8 of 229 cases (3.5%). Of the 229 cases, the DCIS grades were as follows: low in 29 cases (13%), intermediate in 79 cases (36%), and high in 121 cases (53%). Of the 229 cases, necrosis was present in 152 (66.4%) and was comedo necrosis in 99 cases (43%). Of the 229 cases of IC, the tumor stage was as follows: microIC in 36 (16%), T1a in 119 (52%), T1b in 35 (15%), T1c in 28 (12%), T2 in 8 (3%), and T3 in 3 cases (1%). Axillary lymph nodes were staged in 167 patients as follows: N0, 141 cases (84%); N0(i+), 14 cases (8%); and N1, 12 cases (7%). The 12 N1 cases were subclassified by T stage as follows: T1a, 1 case (8%); T1b, 4 cases (33%); T1c, 2 cases (17%); T2, 4 cases (33%); and T3, 1 case (8%). The IC cases of stage upgrading were predominantly smaller than 2 cm (218 of 229; 95%), and more than two-thirds were smaller than 0.5 cm (155 of 229; 95%), most of which were accompanied by extensive DCIS. CONCLUSIONS.­: Approximately one-half of the upgrades were associated with high-grade DCIS, especially with comedo necrosis; nevertheless, the other one-half of the upgrades were due to low- and intermediate-grade DCIS and should not be underestimated. There were few positive results from axillary lymph node biopsies, but they occurred in 3% (7 of 218) of the carcinomas smaller than 2 cm. Our findings indicate that caution is needed when DCIS cases diagnosed by core biopsy are treated nonsurgically with surveillance (with or without neoadjuvant endocrine therapy), given the number of cases (229 of 2943; 8%) that are upgraded to IC and those with axillary lymph node metastases (12 of 167; 7%).

Evaluation of Biomarkers in Multiple Ipsilateral Synchronous Invasive Breast Carcinomas.

CONTEXT.­: The College of American Pathologists guidelines recommend testing additional tumor foci in multifocal invasive breast carcinomas for the biomarkers estrogen receptor (ER), progesterone receptor, and HER2 only if the carcinomas show different morphologies or grades. OBJECTIVE.­: To assess clinical significance of testing for biomarkers in additional tumor foci in multifocal invasive breast tumors. DESIGN.­: Retrospective analysis of 118 patients diagnosed with ipsilateral synchronous multifocal breast carcinomas from January 2015 through March 2016 at Mount Sinai Hospital (New York, New York). RESULTS.­: Eighty-six cases were tested for at least 1 of the 3 biomarkers in additional tumor foci. Fifteen cases (17%) showed discordant staining between the 2 foci for at least one biomarker. Of the 7 of 67 ER-discordant cases (10%), 4 (57%) showed major variation from negative to positive expression, including 3 cases in which a smaller tumor focus was strongly positive for ER whereas the index tumor was negative. Similarly, within the 7 of 67 progesterone receptor-discordant cases (10%), 4 (57%) showed major variation from negative to positive, and in 3 cases with major discordance, the index tumor was negative for progesterone receptor, whereas a smaller focus was positive. A difference in HER2 expression was noted in 5 of 86 cases (6%). In only 5 of the 15 patients (33%) with discordant results, biomarker testing on additional foci would have been offered per the College of American Pathologists recommendations because of differences in histology or grading. Of the remaining 10 patients, 7 (70%) with positive results on smaller foci would have been deprived of appropriate adjuvant systemic treatment if the smaller focus had not been tested. CONCLUSIONS.­: We propose that negative values expressed in the primary tumor be repeated routinely on additional ipsilateral synchronous tumors.