Biopathologic Characterization and Grade Assessment of Breast Cancer With 3-D Multiparametric Ultrasound Combining Shear Wave Elastography and Backscatter Tensor Imaging.

OBJECTIVE: Despite recent improvements in medical imaging, the final diagnosis and biopathologic characterization of breast cancers currently still requires biopsies. Ultrasound is commonly used for clinical examination of breast masses. B-Mode and shear wave elastography (SWE) are already widely used to detect suspicious masses and differentiate benign lesions from cancers. But additional ultrasound modalities such as backscatter tensor imaging (BTI) could provide relevant biomarkers related to tissue organization. Here we describe a 3-D multiparametric ultrasound approach applied to breast carcinomas in the aims of (i) validating the ability of BTI to reveal the underlying organization of collagen fibers and (ii) assessing the complementarity of SWE and BTI to reveal biopathologic features of diagnostic interest. METHODS: Three-dimensional SWE and BTI were performed ex vivo on 64 human breast carcinoma samples using a linear ultrasound probe moved by a set of motors. Here we describe a 3-D multiparametric representation of the breast masses and quantitative measurements combining B-mode, SWE and BTI. RESULTS: Our results reveal for the first time that BTI can capture the orientation of the collagen fibers around tumors. BTI was found to be a relevant marker for assessing cancer stages, revealing a more tangent tissue orientation for in situ carcinomas than for invasive cancers. In invasive cases, the combination of BTI and SWE parameters allowed for classification of invasive tumors with respect to their grade with an accuracy of 95.7%. CONCLUSION: Our results highlight the potential of 3-D multiparametric ultrasound imaging for biopathologic characterization of breast tumors.

Proof of Concept of 3-D Backscatter Tensor Imaging Tomography for Non-invasive Assessment of Human Breast Cancer Collagen Organization.

Tumor growth, similarly to several other pathologies, tends to change the structural orientation of soft tissue fibers, which can become relevant markers for diagnosis. Current diagnosis protocols may require a biopsy for histological analysis, which is an invasive, painful and stressful procedure with a minimum turnaround time of 2 d. Otherwise, diagnosis may involve the use of complex methods with limited availability such as diffusion tensor imaging (magnetic resonance diffusion tensor imaging), which is not widely used in medical practice. Conversely, advanced methodologies in ultrasound imaging such as backscatter tensor imaging (BTI) might become a routine procedure in clinical practice at a limited cost. This method evaluates the local organization of soft tissues based on the spatial coherence of their backscattered ultrasonic echoes. Previous work has proven that BTI applied with matrix probes enables measurement of the orientation of soft tissue fibers, especially in the myocardium. The aims of the study described here were (i) to present for the first time a methodology for performing BTI in a volume on ex vivo human breast tumors using a linear probe and (ii) to display a first proof of concept of the link between BTI measurements and the orientation of collagen fibers.

Assessment of the Molecular Heterogeneity of E-Cadherin Expression in Invasive Lobular Breast Cancer.

Mutations and loss of E-cadherin protein expression define the vast majority of invasive lobular carcinomas. In a subset of these cases, the heterogeneous expression of E-cadherin is observed either as wild-type (strong membranous) expression or aberrant expression (cytoplasmic expression). However, it is unclear as to whether the two components would be driven by distinct genetic or epigenetic alterations. Here, we used whole genome DNA sequencing and methylation array profiling of two separately dissected components of nine invasive lobular carcinomas with heterogeneous E-cadherin expression. E-cadherin negative and aberrant/positive components of E-cadherin heterogeneous tumours showed a similar mutational, copy number and promoter methylation repertoire, suggesting they arise from a common ancestor, as opposed to the collision of two independent tumours. We found that the majority of E-cadherin heterogeneous tumours harboured CDH1 mutations in both the E-cadherin negative and aberrant/positive components together with somatic mutations in additional driver genes known to be enriched in both pure invasive carcinomas of no special type and invasive lobular breast cancers, whereas these were less commonly observed in CDH1 wild-type tumours. CDH1 mutant tumours also exhibited a higher mutation burden as well as increased presence of APOBEC-dependent mutational signatures 2 and 13 compared to CDH1 wild-type tumours. Together, our results suggest that regardless of E-cadherin protein expression, tumours showing heterogeneous expression of E-cadherin should be considered as part of the spectrum of invasive lobular breast cancers.

[2021 update of the GEFPICS' recommendations for HER2 status assessment in invasive breast cancer in France]. / Mise à jour 2021 des recommandations du GEFPICS pour l'évaluation du statut HER2 dans les cancers infiltrants du sein en France.

The last international guidelines on HER2 determination in breast cancer have been updated in 2018 by the American Society of Clinical Oncology and College of American Pathologists, on the basis of a twenty-year practice and results of numerous clinical trials. Moreover, the emerging HER2-low concept for 1+ and 2+ non amplified breast cancers lead to refine French practices for HER2 status assessment. The GEFPICS group, composed of expert pathologists, herein presents the latest French recommendations for HER2 status evaluation in breast cancer, taking into account the ASCO/CAP guidelines and introducing the HER2-low concept. In the era of personalized medicine, HER2 status assessment remains one of the most important biomarkers in breast cancer and its quality guaranties the optimal patients' care. French pathologists' commitment in theranostic biomarker quality is more than ever required to provide the most efficient cares in oncology.

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.

The path to a better biomarker: application of a risk management framework for the implementation of PD-L1 and TILs as immuno-oncology biomarkers in breast cancer clinical trials and daily practice.

Immune checkpoint inhibitor therapies targeting PD-1/PD-L1 are now the standard of care in oncology across several hematologic and solid tumor types, including triple negative breast cancer (TNBC). Patients with metastatic or locally advanced TNBC with PD-L1 expression on immune cells occupying ≥1% of tumor area demonstrated survival benefit with the addition of atezolizumab to nab-paclitaxel. However, concerns regarding variability between immunohistochemical PD-L1 assay performance and inter-reader reproducibility have been raised. High tumor-infiltrating lymphocytes (TILs) have also been associated with response to PD-1/PD-L1 inhibitors in patients with breast cancer (BC). TILs can be easily assessed on hematoxylin and eosin-stained slides and have shown reliable inter-reader reproducibility. As an established prognostic factor in early stage TNBC, TILs are soon anticipated to be reported in daily practice in many pathology laboratories worldwide. Because TILs and PD-L1 are parts of an immunological spectrum in BC, we propose the systematic implementation of combined PD-L1 and TIL analyses as a more comprehensive immuno-oncological biomarker for patient selection for PD-1/PD-L1 inhibition-based therapy in patients with BC. Although practical and regulatory considerations differ by jurisdiction, the pathology community has the responsibility to patients to implement assays that lead to optimal patient selection. We propose herewith a risk-management framework that may help mitigate the risks of suboptimal patient selection for immuno-therapeutic approaches in clinical trials and daily practice based on combined TILs/PD-L1 assessment in BC. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

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.

Medico-economic impact of MSKCC non-sentinel node prediction nomogram for ER-positive HER2-negative breast cancers.

BACKGROUND: Avoiding axillary lymph node dissection (ALND) for invasive breast cancers with isolated tumor cells or micrometastatic sentinel node biopsy (SNB) could decrease morbidity with minimal clinical significance. PURPOSE: The aim of this study is to simulate the medico-economic impact of the routine use of the MSKCC non-sentinel node (NSN) prediction nomogram for ER+ HER2- breast cancer patients. METHODS: We studied 1036 ER+ HER2- breast cancer patients with a metastatic SNB. All had a complementary ALND. For each patient, we calculated the probability of the NSN positivity using the MSKCC nomogram. After validation of this nomogram in the population, we described how the patients' characteristics spread as the threshold value changed. Then, we performed an economic simulation study to estimate the total cost of caring for patients treated according to the MSKCC predictive nomogram results. RESULTS: A 0.3 threshold discriminate the type of sentinel node (SN) metastases: 98.8% of patients with pN0(i+) and 91.6% of patients with pN1(mic) had a MSKCC score under 0.3 (false negative rate = 6.4%). If we use the 0.3 threshold for economic simulation, 43% of ALND could be avoided, reducing the costs of caring by 1 051 980 EUROS among the 1036 patients. CONCLUSION: We demonstrated the cost-effectiveness of using the MSKCC NSN prediction nomogram by avoiding ALND for the pN0(i+) or pN1(mic) ER+ HER2- breast cancer patients with a MSKCC score of less than or equal to 0.3.

[Pre-analytical stage for biomarker assessment in breast cancer: 2014 update of the GEFPICS' guidelines in France]. / Recommandations du GEFPICS concernant la phase pré-analytique pour l'évaluation de HER2 et des récepteurs hormonaux dans le cancer du sein : mise à jour 2014.

Biomarker assessment of breast cancer tumor samples is part of the routine workflow of pathology laboratories. International guidelines have recently been updated, with special regards to the pre-analytical steps that are critical for the quality of immunohistochemical and in situ hybridization procedures, whatever the biomarker analyzed. Fixation and specimen handling protocols must be standardized, validated and carefully tracked. Cooperation and training of the personnel involved in the specimen workflow (e.g. radiologists, surgeons, nurses, technicians and pathologists) are of paramount importance. The GEFPICS' update of the recommendations herein details and comments the different steps of the pre-analytical process. Application of these guidelines and participation to quality insurance programs are mandatory to ensure the correct evaluation of oncotheranostic biomarkers.

Confrontation of immunohistochemistry and fluorescent in situ hybridization for the assessment of HER-2/ neu (c-erbb-2) status in urothelial carcinoma.

Specific treatments targeted toward oncogenes expressed in cancer cells are currently under development. Patients with urothelial carcinomas showing HER-2/ neu (human epidermal growth factor receptor 2) overexpression are candidates for such a specific treatment (trastuzumab). However, to be effective, this therapeutic approach requires an extremely reliable evaluation of HER-2/ neu status in tumors. In order to assess the status of expression of this gene and to optimize its assessment, we analyzed a series of 64 primary urothelial carcinomas using immunohistochemistry (IHC) with the CB11 monoclonal antibody coupled with fluorescent in situ hybridization (FISH) in 21 cases. Strong HER-2/ neu overexpression was detected using IHC in 15 of the 64 (23%) cases analyzed, and this rate rose to 33% for patients with metastases. HER-2/ neu overexpression, as revealed using IHC, is strongly associated (95%) with gene amplification assessed using FISH. Patients with urothelial carcinomas overexpressing HER-2/ neu using IHC are potential candidates for targeted chemotherapy.

Histological margin assessment for breast ductal carcinoma in situ: precision and implications.

Local recurrence after lumpectomy for ductal carcinoma in situ (DCIS) is a major concern and is related to residual disease in the breast. We studied the predictive value of lumpectomy margins for residual DCIS and compared our results and pathological processing techniques with those published in the literature. Margin status was determined for 89 patients with screen-detected DCIS who had lumpectomy and re-excision, for the presence and extent of residual disease. Margin width was defined as the narrowest distance between tumor and any inked margin or, where margins were positive, classified into focal involvement (<1 mm of the inked surface involved), minimal (>or=1<15 mm) and extensive (>or=15 mm). The amount of residual tumor was quantified according to the number of ducts involved with tumor: small (fewer than 10 ducts) or large (10 or more ducts) residuum. The initial margin status was a significant predictor for the presence of residual tumor in re-excision specimens (P=0.006). There was residual tumor in 44 and 45% of close non-involved (>1 and 25 mm,respectively, showing residual disease. The presence of residual tumor was not significantly related to age, mammographic appearance, nuclear grade or intraductal necrosis. The initial margin status was found to predict for the amount of residual tumor. With careful margin assessment, margin status after lumpectomy for DCIS can be used to predict for the presence and amount of residual tumor in the breast and is a guide to further management decisions. A standard for margin status reporting and pathological processing of screen-detected DCIS in situ lesions will help in the interpretation of data from different institutions.

HER2 status in patients with breast carcinoma is not modified selectively by preoperative chemotherapy and is stable during the metastatic process.

BACKGROUND: The objective of this study was to determine whether HER2 expression levels in breast carcinomas were modified by chemotherapy or during the metastatic process. METHODS: HER2 expression was analyzed on sequential tissue specimens taken from the primary tumor before patients received preoperative chemotherapy (CT) and from post-CT residual breast tumor or at a metastatic site. The first group of patients included 59 women who presented with T2-T4,N1-N2 breast carcinoma and were treated by preoperative anthracycline-based CT and then underwent surgery. The second group included 44 patients with metastatic breast carcinoma localized to the lung (27 patients) or to the liver (17 patients). HER2 status was determined by immunohistochemistry using an anti-p185(HER/neu) monoclonal antibody and was classified as overexpressed or not overexpressed. RESULTS: Among the patients who received preoperative CT, HER2 overexpression was observed in 15 of 59 patients (25%). A complete pathologic response was observed in 2 of these 15 patients. HER2 still was overexpressed in 11 of 13 remaining residual tumors and was no longer detectable in 2 tumors. In addition, the 29 tumors with no HER2 overexpression before CT remained negative after treatment. In patients with metastatic breast carcinoma, HER2 was overexpressed in 11 of 44 primary tumors (25%). In 9 of these 11 tumors, HER2 overexpression was maintained in the metastases (9 pulmonary metastases and 4 hepatic metastases). In two patients who had low levels of HER2 overexpression in their primary tumors, no staining was observed in the secondary tumor (one pulmonary tumor and one liver tumor). There were no tumors in which the overexpression of HER2 was found only in the metastasis. CONCLUSIONS: The current study showed that, in most patients, HER2 overexpression was unchanged after CT and in metastatic sites. No HER2 negative primary tumors became HER2 positive after patients received CT or during the metastatic process. In a few patients, a diminution in the level of HER2 expression was observed after CT or in secondary tumors. This may have been due to a transitory state of altered tumor cells or to the selection of HER2 negative tumor cells clones.