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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 hocsTILs 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.

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 hocsTILs 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.

Les dernières recommandations internationales pour l’évaluation du statut HER2 dans les cancers du sein, basées sur vingt ans d’expérience et sur les résultats de nombreuses études cliniques, ont vu le jour en 2018. D’autre part, la notion émergente de catégorie HER2 faible pour les cancers du sein de score 1+ en immunohistochimie ou de score 2+ sans amplification du gène HER2 en hybridation in situinvite à adapter les pratiques des pathologistes français. Ces modifications importantes et les bouleversements à venir sont l’occasion, pour le GEFPICS, de proposer une mise à jour des recommandations françaises pour l’évaluation du statut HER2 dans les cancers du sein. À l’ère de la médecine personnalisée, la détermination du statut HER2 reste un élément phare dans le panel des biomarqueurs théranostiques des cancers du sein, et sa bonne évaluation est garante d’une prise en charge optimale des patientes atteintes de cancer du sein. L’implication des pathologistes français dans la qualité des tests théranostiques témoigne de leur rôle essentiel dans la prise en charge des patients en cancérologie.

Primary triple-negative invasive lobular breast carcinomas (TN-ILCs), which do not express hormone receptors and HER2 at diagnosis, are rare and poorly known. In this study, we analyzed the largest TN-ILC series ever reported in the literature, in comparison to phenotypically similar breast tumor subtypes: triple-negative invasive ductal carcinoma (TN-IDC) and hormone receptor-positive invasive lobular carcinoma (HR + ILC). All primary TN-ILCs registered in our database between 2000 and 2018 (n= 38) were compared to tumors from control groups, matched by stage and Elston/Ellis grade, with regard to clinical, pathologic, and immunohistochemical characteristics. A comparative molecular analysis (whole-exome and RNA sequencing using next-generation technology) was also performed. We found that TN-ILC patients were older than those with HR + ILC (P= 0.002) or TN-IDC (P< 0.001). Morphologically, TN-ILCs had aggressive phenotypes, with more pleomorphism (P= 0.003) and higher nuclear grades than HR + ILCs (P= 0.009). Immunohistochemistry showed that TN-ILCs less frequently expressed basal markers (CK5/6, EGFR and SOX10) than TN-IDCs (P< 0.001), while androgen receptor (AR) positivity was more prevalent (P< 0.001). Survival curves analysis did not show differences between TN-ILC and TN-IDC patients, while overall and distant metastasis-free survival were significantly worse compared to those with HR + ILCs (P= 0.047 and P= 0.039, respectively). At a molecular level, we found that TN-ILCs had particular transcriptomic profiles, characterized by increased AR signaling, and associated with frequent alterations in the PI3K network and ERBB2. Interestingly, whole-exome analysis also identified three specific recurrent ESRRAhotspot mutations in these tumors, which have never been described in breast cancer to date and which were absent in the other two tumor subtypes. Our findings highlight that TN-ILC is a unique aggressive breast cancer associated with elderly age, which belong to the luminal androgen receptor subtype as determined by immunohistochemistry and transcriptomic profiling. Moreover, it harbors specific molecular alterations (PI3K, ERBB2and ESRRA) which may pave the way for new targeted therapeutic strategies.

Primary triple-negative invasive lobular breast carcinomas (TN-ILCs), which do not express hormone receptors and HER2 at diagnosis, are rare and poorly known. In this study, we analyzed the largest TN-ILC series ever reported in the literature, in comparison to phenotypically similar breast tumor subtypes: triple-negative invasive ductal carcinoma (TN-IDC) and hormone receptor-positive invasive lobular carcinoma (HR + ILC). All primary TN-ILCs registered in our database between 2000 and 2018 (n= 38) were compared to tumors from control groups, matched by stage and Elston/Ellis grade, with regard to clinical, pathologic, and immunohistochemical characteristics. A comparative molecular analysis (whole-exome and RNA sequencing using next-generation technology) was also performed. We found that TN-ILC patients were older than those with HR + ILC (P= 0.002) or TN-IDC (P< 0.001). Morphologically, TN-ILCs had aggressive phenotypes, with more pleomorphism (P= 0.003) and higher nuclear grades than HR + ILCs (P= 0.009). Immunohistochemistry showed that TN-ILCs less frequently expressed basal markers (CK5/6, EGFR and SOX10) than TN-IDCs (P< 0.001), while androgen receptor (AR) positivity was more prevalent (P< 0.001). Survival curves analysis did not show differences between TN-ILC and TN-IDC patients, while overall and distant metastasis-free survival were significantly worse compared to those with HR + ILCs (P= 0.047 and P= 0.039, respectively). At a molecular level, we found that TN-ILCs had particular transcriptomic profiles, characterized by increased AR signaling, and associated with frequent alterations in the PI3K network and ERBB2. Interestingly, whole-exome analysis also identified three specific recurrent ESRRAhotspot mutations in these tumors, which have never been described in breast cancer to date and which were absent in the other two tumor subtypes. Our findings highlight that TN-ILC is a unique aggressive breast cancer associated with elderly age, which belong to the luminal androgen receptor subtype as determined by immunohistochemistry and transcriptomic profiling. Moreover, it harbors specific molecular alterations (PI3K, ERBB2and ESRRA) which may pave the way for new targeted therapeutic strategies.

Adenoid cystic carcinoma (ACC) of the breast with a predominant solid pattern is difficult to diagnose with certainty and differentiate from more common triple-negative breast cancers (TNBCs) of basal-phenotype. To better characterize solid ACC, we performed a clinical, morphological, immunohistochemical, and molecular comparative analysis of 33 ACCs of the breast comprising 17 solid variant ACCs and 16 conventional ACCs. Solid ACCs displayed basaloid morphology with an exclusive or predominant epithelial cell population associated with decreased myoepithelial differentiation, while demonstrating MYB protein overexpression similar to the more common type of ACC. Strong and diffuse MYB expression by immunochemistry was observed in 14/17 (82%) of solid ACCs while MYBrearrangements were detected by break apart fluorescence in situ hybridization (FISH) in only 3/16 (19%) of solid ACCs. Conversely, weak MYB immunohistochemical expression was observed in only 7/204 (3%) of TNBC. Solid ACCs displayed a transcriptomic profile distinct from conventional ACCs with 549 genes showing a highly significant differential expression between conventional and solid ACC [false discovery rate (FDR) < 0.01; log2FC > |1|]. EnrichR and Kegg Pathway analyses identified PI3K-Akt and focal adhesion signaling pathways as significantly overexpressed in conventional ACCs compared with solid ACCs which significantly overexpressed the nitrogen metabolism pathway. CREBBPmutations and NOTCHactivating gene mutations were only present in solid ACCs, concerning 5/16 (31%) of cases for each gene. Tumors with NOTCHactivating mutations displayed a strong diffuse nuclear NICD1 staining, an established marker of Notch pathway activation. Solid ACCs also differed from basal-type TNBC, with fewer TP53mutations and a more stable genomic profile on array comparative genomic hybridization (CGH). In summary, solid-type ACC of the breast is a distinct molecular entity within the ACC family and is different from common basal-type TNBC. MYB is a diagnostically useful biomarker of solid ACC and NOTCHcould be a novel potential therapeutic target in 30% of cases.

Adenoid cystic carcinoma (ACC) of the breast with a predominant solid pattern is difficult to diagnose with certainty and differentiate from more common triple-negative breast cancers (TNBCs) of basal-phenotype. To better characterize solid ACC, we performed a clinical, morphological, immunohistochemical, and molecular comparative analysis of 33 ACCs of the breast comprising 17 solid variant ACCs and 16 conventional ACCs. Solid ACCs displayed basaloid morphology with an exclusive or predominant epithelial cell population associated with decreased myoepithelial differentiation, while demonstrating MYB protein overexpression similar to the more common type of ACC. Strong and diffuse MYB expression by immunochemistry was observed in 14/17 (82%) of solid ACCs while MYBrearrangements were detected by break apart fluorescence in situ hybridization (FISH) in only 3/16 (19%) of solid ACCs. Conversely, weak MYB immunohistochemical expression was observed in only 7/204 (3%) of TNBC. Solid ACCs displayed a transcriptomic profile distinct from conventional ACCs with 549 genes showing a highly significant differential expression between conventional and solid ACC [false discovery rate (FDR) < 0.01; log2FC > |1|]. EnrichR and Kegg Pathway analyses identified PI3K-Akt and focal adhesion signaling pathways as significantly overexpressed in conventional ACCs compared with solid ACCs which significantly overexpressed the nitrogen metabolism pathway. CREBBPmutations and NOTCHactivating gene mutations were only present in solid ACCs, concerning 5/16 (31%) of cases for each gene. Tumors with NOTCHactivating mutations displayed a strong diffuse nuclear NICD1 staining, an established marker of Notch pathway activation. Solid ACCs also differed from basal-type TNBC, with fewer TP53mutations and a more stable genomic profile on array comparative genomic hybridization (CGH). In summary, solid-type ACC of the breast is a distinct molecular entity within the ACC family and is different from common basal-type TNBC. MYB is a diagnostically useful biomarker of solid ACC and NOTCHcould be a novel potential therapeutic target in 30% of cases.

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.

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.

La chimiothérapie néoadjuvante est une option thérapeutique de plus en plus utilisée dans la stratégie de prise en charge des cancers du sein. La réponse histologique au traitement néoadjuvant, essentiellement sous la forme d’une réponse pathologique complète, est aujourd’hui utilisée comme facteur pronostique et comme critère validé d’évaluation de l’efficacité d’un traitement dans les essais cliniques évaluant de nouvelles stratégies thérapeutiques. Il existe, cependant, encore aujourd’hui un manque de standardisation dans la prise en charge des prélèvements mammaires dans le cadre d’un traitement néoadjuvant (manque d’uniformité dans la prise en charge macroscopique, dans l’interprétation microscopique, dans les données présentes dans le compte rendu de pathologie et dans la définition même de la « réponse pathologique complète »), cette hétérogénéité de prise en charge pouvant entraîner des biais importants dans l’évaluation du pronostic de la patiente ou dans l’analyse des résultats des études cliniques. Le GEFPICS propose donc des recommandations pour la prise en charge des prélèvements mammaires et ganglionnaires avant traitement (prise en charge des biopsies, données minimales à fournir dans le compte rendu de pathologie) et après traitement (prise en charge macroscopique, interprétation microscopique, classifications post-thérapeutiques).

Triple-negative breast cancer (TNBC) patients have an increased risk of developing visceral metastases and other primary nonbreast cancers, particularly lung cancer. The differential diagnosis of TNBC metastases and primary cancers from other organs can be difficult due to lack of a TNBC standard immunoprofile. We analyzed the diagnostic value of estrogen receptor, progesterone receptor, human epidermal growth factor receptor, thyroid transcription factor-1 (TTF1), Napsin A, mammaglobin, gross cystic disease fluid protein 15 (GCDFP15), Sry-related HMg-Box gene 10 (SOX10), GATA-binding protein 3 (GATA3), and androgen receptor in a series of 207 TNBC and 152 primary lung adenocarcinomas (LA). All tested TNBCs were TTF1 and Napsin A-negative. When comparing TNBC and TTF1-positive or negative LA, SOX10 had the best sensitivity (62.3%) and specificity (100%) as a marker in favor of TNBC compared with LA, irrespective of TTF1 status (P<0.0001). GATA3 had moderate sensitivity (30.4%) and excellent specificity (98.7%) and misclassified only 2/152 LA (1.3%). GCDFP15 had a moderate sensitivity (20.8%) and excellent specificity (98%) and misclassified only 3/152 (2%) LA. Mammaglobin and androgen receptor had moderate sensitivities (38.2% and 30%), good specificities (81.6% and 86%), and misclassified 28/152 and 21/152 LAs, respectively. In multivariate analysis, the best markers, enabling the distinction between SOX10-negative TNBC and TTF1 and Napsin A-negative LA were GATA3 (odds ratio=33.5; 95% confidence interval, 7.3-153.5; P<0.0001) and GCDFP15 (odds ratio=31.7; 95% confidence interval, 6.9-145.6; P<0.0001). Only 13/207 (6.3%) TNBC cases did not express any aforementioned marker. On the basis of our results, the best sequential immunohistochemical analysis to differentiate TNBC from TTF1-negative LA is first SOX10 followed by GATA3, and finally GCDFP15. This order is important in the diagnostic workup of small biopsies from lung nodules in women with a previous history of TNBC.

Dermatofibrosarcoma protuberans is underlined by recurrent collagen type I alpha 1 chain-platelet-derived growth factor B chain(COL1A1-PDGFB) fusions but ~ 4% of typical dermatofibrosarcoma protuberans remain negative for this translocation in routine molecular screening. We investigated a series of 21 cases not associated with the pathognomonic COL1A1-PDGFBfusion on routine fluorescence in situ hybridization (FISH) testing. All cases displayed morphological and clinical features consistent with the diagnosis of dermatofibrosarcoma protuberans. RNA-sequencing analysis was successful in 20 cases. The classical COL1A1-PDGFBfusion was present in 40% of cases (n= 8/20), and subsequently confirmed with a COL1A1break-apart FISH probe in all but one case (n= 7/8). 55% of cases (n= 11/20) displayed novel PDGFDrearrangements; PDGFDbeing fused either to the 5′ part of COL6A3(2q37.3) (n= 9/11) or EMILIN2(18p11) (n= 2/11). All rearrangements led to in-frame fusion transcripts and were confirmed at genomic level by FISH and/or array-comparative genomic hybridization. PDGFD-rearranged dermatofibrosarcoma protuberans presented clinical outcomes similar to typical dermatofibrosarcoma protuberans. Notably, the two EMILIN2-PDGFDcases displayed fibrosarcomatous transformation and homozygous deletions of CDKN2Aat genomic level. We report the first recurrent molecular variant of dermatofibrosarcoma protuberans involving PDGFD, which functionally mimic bona fide COL1A1-PDGFB fusions, leading presumably to a similar autocrine loop-stimulating PDGFRB. This study also emphasizes that COL1A1-PDGFBfusions can be cytogenetically cryptic on FISH testing in a subset of cases, thereby representing a diagnostic pitfall that pathologists should be aware of.

Medullary breast carcinoma (MBC) is a rare subtype of triple-negative breast cancer with specific genomic features within the spectrum of basal-like carcinoma (BLC). In this study of 19 MBCs and 36 non-MBC BLCs, we refined the transcriptomic and genomic knowledge about this entity. Unsupervised and supervised analysis of transcriptomic profiles confirmed that MBC clearly differs from non-MBC BLC, with 92 genes overexpressed and 154 genes underexpressed in MBC compared with non-MBC BLC. Immunity-related pathways are the most differentially represented pathways in MBC compared with non-MBC BLC. The proapoptotic gene BCLG(official name BCL2L14) is by far the most intensely overexpressed gene in MBC. A quantitative RT-PCR validation study conducted in 526 breast tumors corresponding to all molecular subtypes documented the specificity of BCLGoverexpression in MBC, which was confirmed at the protein level by immunohistochemistry. We also found that most MBCs belong to the immunomodulatory triple-negative breast cancer subtype. Using pan-genomic analysis, it was found that MBC harbors more losses of heterozygosity than non-MBC BLC. These observations corroborate the notion that MBC remains a distinct entity that could benefit from specific treatment strategies (such as deescalation or targeted therapy) adapted to this rare tumor type.

Solid papillary carcinoma with reverse polarity is a rare breast cancer of favorable prognosis that can be difficult to diagnose. We report here nine additional cases of this tumor, and we describe its morphologic, immunohistochemical and molecular profile in comparison to other types of papillary and micropapillary lesions of the breast that are intraductal papilloma with usual ductal hyperplasia, encapsulated papillary carcinoma, solid papillary carcinoma and invasive micropapillary carcinoma. We studied nine cases of this special papillary tumor and six of each other types mentioned above. We found that solid papillary carcinoma with reverse polarity harbor specific morphologic features as cuboid or tall cells with abundant eosinophilic cytoplasms located at the basal pole giving the impression of reverse nuclear polarity. Nuclei were sometimes grooved. Immunohistochemistry demonstrated the lack of myoepithelial cells, as in encapsulated papillary carcinoma and solid papillary carcinoma, questioning their invasive nature. Seven of nine solid papillary carcinoma with reverse polarity showed a low Ki67 proliferative index (Ki67 <5%). They showed expression of CK5/6 as in intraductal papilloma with usual ductal hyperplasia. They showed expression of calretinin and a low or lack of hormonal receptor (HR) expression that were not observed in other breast tumors studied. By whole-exome analysis, seven of nine solid papillary carcinomas with reverse polarity (78%) harbored a hotspot mutation in IDH2 (R172) that was totally absent in other groups. Six of nine tumors (67%) also harbored PRUNE2 mutation, including the two IDH2 wild-type cases. We also demonstrated for the first time in this breast tumor, immunostaining with a specific antibody IDH1/2 mutant R132/R172 (7/9) that can highlight IDH2 mutation. Moreover, transcriptomic analysis showed that proteoglycan pathway was significantly enriched. Our findings support the fact that solid papillary carcinoma with reverse polarity is a singular breast neoplasm that can be distinguished from other papillary breast tumors.

Solid papillary carcinoma with reverse polarity is a rare breast cancer of favorable prognosis that can be difficult to diagnose. We report here nine additional cases of this tumor, and we describe its morphologic, immunohistochemical and molecular profile in comparison to other types of papillary and micropapillary lesions of the breast that are intraductal papilloma with usual ductal hyperplasia, encapsulated papillary carcinoma, solid papillary carcinoma and invasive micropapillary carcinoma. We studied nine cases of this special papillary tumor and six of each other types mentioned above. We found that solid papillary carcinoma with reverse polarity harbor specific morphologic features as cuboid or tall cells with abundant eosinophilic cytoplasms located at the basal pole giving the impression of reverse nuclear polarity. Nuclei were sometimes grooved. Immunohistochemistry demonstrated the lack of myoepithelial cells, as in encapsulated papillary carcinoma and solid papillary carcinoma, questioning their invasive nature. Seven of nine solid papillary carcinoma with reverse polarity showed a low Ki67 proliferative index (Ki67 <5%). They showed expression of CK5/6 as in intraductal papilloma with usual ductal hyperplasia. They showed expression of calretinin and a low or lack of hormonal receptor (HR) expression that were not observed in other breast tumors studied. By whole-exome analysis, seven of nine solid papillary carcinomas with reverse polarity (78%) harbored a hotspot mutation in IDH2 (R172) that was totally absent in other groups. Six of nine tumors (67%) also harbored PRUNE2 mutation, including the two IDH2 wild-type cases. We also demonstrated for the first time in this breast tumor, immunostaining with a specific antibody IDH1/2 mutant R132/R172 (7/9) that can highlight IDH2 mutation. Moreover, transcriptomic analysis showed that proteoglycan pathway was significantly enriched. Our findings support the fact that solid papillary carcinoma with reverse polarity is a singular breast neoplasm that can be distinguished from other papillary breast tumors.

The failure to accurately define tumor margins during breast conserving surgery (BCS) results in a 20% re-excision rate. The present paper reports the investigation to evaluate the potential of terahertz imaging for breast tissue recognition within the under-explored 300 – 600 GHz range. Such a frequency window matches new BiCMOS technology capabilities and thus opens up the opportunity for near-field terahertz imaging using these devices. To assess the efficacy of this frequency band, data from 16 freshly excised breast tissue samples were collected and analyzed directly after excision. Complex refractive indices have been extracted over the as-mentioned frequency band, and amplitude frequency images show some contrast between tissue types. Principal component analysis (PCA) has also been applied to the data in an attempt to automate tissue classification. Our observations suggest that the dielectric response could potentially provide contrast for breast tissue recognition within the 300 – 600 GHz range. These results open the way for silicon-based terahertz subwavelength near field imager design, efficient up to 600 GHz to address ex vivo life-science applications.

Le dépistage mammographique s’accompagne d’un surdiagnostic, inhérent à l’acte de dépistage mais lié aussi, dans un certain nombre de cas, à une surinterprétation des images mammographiques, de la part des radiologues, et des lésions frontières de la part les pathologistes. Les lésions frontières en pathologie mammaire sont des néoplasies intraépithéliales dont le diagnostic est difficile du fait de l’existence d’un continuum lésionnel morphologique, avec des limites floues d’une catégorie lésionnelle à l’autre, diminuant la reproductibilité diagnostique interobservateur. L’Institut national du cancer a récemment proposé un plan d’action pour la rénovation du dépistage organisé en France et l’amélioration de la qualité en pathologie. Dans le cadre de ce plan une expérimentation de double lecture des lésions frontières est proposée, mais ses modalités pratiques restent à définir.

The diagnosis of a uterine smooth muscle lesion is, in the majority of cases, straightforward. However, in a small number of cases, the morphological criteria used in such lesions cannot differentiate with certainty a benign from a malignant lesion and a diagnosis of smooth muscle tumor with uncertain malignant potential (STUMP) is made. Uterine leiomyosarcomas are often easy to diagnose but it is difficult or even impossible to identify a prognostic factor at the moment of the diagnosis with the exception of the stage. We hypothesize, for uterine smooth muscle lesions, that there is a gradient of genomic complexity that correlates to outcome. We first tested this hypothesis on STUMP lesions in a previous study and demonstrated that this 'gray category' could be split according to genomic index into two groups. A benign group, with a low to moderate alteration rate without recurrence and a malignant group, with a highly rearranged profile akin to uterine leiomyosarcomas. Here, we analyzed a large series of 77 uterine smooth muscle lesions (from 76 patients) morphologically classified as 19 leiomyomas, 14 STUMP and 44 leiomyosarcomas with clinicopathological and genomic correlations. We confirmed that genomic index with a cut-off=10 is a predictor of recurrence (P<0.0001) and with a cut-off=35 is a marker for poor overall survival (P=0.035). For the tumors confined to the uterus, stage as a prognostic factor was not useful in survival prediction. At stage I, among the tumors reclassified as molecular leiomyosarcomas (ie, genomic index ≥10), the poor prognostic markers were: 5p gain (overall survival P=0.0008), genomic index at cut-off=35 (overall survival P=0.0193), 13p loss including RB1(overall survival P=0.0096) and 17p gain including MYOCDgain (overall survival P=0.0425). Based on these findings (and the feasibility of genomic profiling by array-comparative genomic hybridization), genomic index, 5p and 17p gains prognostic value could be evaluated in future prospective chemotherapy trials.

The diagnosis of a uterine smooth muscle lesion is, in the majority of cases, straightforward. However, in a small number of cases, the morphological criteria used in such lesions cannot differentiate with certainty a benign from a malignant lesion and a diagnosis of smooth muscle tumor with uncertain malignant potential (STUMP) is made. Uterine leiomyosarcomas are often easy to diagnose but it is difficult or even impossible to identify a prognostic factor at the moment of the diagnosis with the exception of the stage. We hypothesize, for uterine smooth muscle lesions, that there is a gradient of genomic complexity that correlates to outcome. We first tested this hypothesis on STUMP lesions in a previous study and demonstrated that this 'gray category' could be split according to genomic index into two groups. A benign group, with a low to moderate alteration rate without recurrence and a malignant group, with a highly rearranged profile akin to uterine leiomyosarcomas. Here, we analyzed a large series of 77 uterine smooth muscle lesions (from 76 patients) morphologically classified as 19 leiomyomas, 14 STUMP and 44 leiomyosarcomas with clinicopathological and genomic correlations. We confirmed that genomic index with a cut-off=10 is a predictor of recurrence (P<0.0001) and with a cut-off=35 is a marker for poor overall survival (P=0.035). For the tumors confined to the uterus, stage as a prognostic factor was not useful in survival prediction. At stage I, among the tumors reclassified as molecular leiomyosarcomas (ie, genomic index ≥10), the poor prognostic markers were: 5p gain (overall survival P=0.0008), genomic index at cut-off=35 (overall survival P=0.0193), 13p loss including RB1(overall survival P=0.0096) and 17p gain including MYOCDgain (overall survival P=0.0425). Based on these findings (and the feasibility of genomic profiling by array-comparative genomic hybridization), genomic index, 5p and 17p gains prognostic value could be evaluated in future prospective chemotherapy trials.

Breast Cancer is one of the most frequently diagnosed cancer diseases worldwide, and the most common invasive tumour for women. As with all cancers, early detection plays a major role in reducing the mortality and morbidity rate. Currently, most breast cancers are detected due to clinical symptoms, or by screening mammography. The limitations of these techniques have resulted in research of alternative methods for imaging and detecting breast cancer. Apart from this, it is essential to define precise tumour margins during breast-conserving surgeries to reduce the re-excision rate. This study presents the advances in the development of a silicon-based THz sub-wavelength imager usable in life science applications, especially for tumour margin identification.

Dermatofibrosarcoma protuberans is underlined by recurrent collagen type I alpha 1 chain-platelet-derived growth factor B chain(COL1A1-PDGFB) fusions but ~ 4% of typical dermatofibrosarcoma protuberans remain negative for this translocation in routine molecular screening. We investigated a series of 21 cases not associated with the pathognomonic COL1A1-PDGFBfusion on routine fluorescence in situ hybridization (FISH) testing. All cases displayed morphological and clinical features consistent with the diagnosis of dermatofibrosarcoma protuberans. RNA-sequencing analysis was successful in 20 cases. The classical COL1A1-PDGFBfusion was present in 40% of cases (n= 8/20), and subsequently confirmed with a COL1A1break-apart FISH probe in all but one case (n= 7/8). 55% of cases (n= 11/20) displayed novel PDGFDrearrangements; PDGFDbeing fused either to the 5′ part of COL6A3(2q37.3) (n= 9/11) or EMILIN2(18p11) (n= 2/11). All rearrangements led to in-frame fusion transcripts and were confirmed at genomic level by FISH and/or array-comparative genomic hybridization. PDGFD-rearranged dermatofibrosarcoma protuberans presented clinical outcomes similar to typical dermatofibrosarcoma protuberans. Notably, the two EMILIN2-PDGFDcases displayed fibrosarcomatous transformation and homozygous deletions of CDKN2Aat genomic level. We report the first recurrent molecular variant of dermatofibrosarcoma protuberans involving PDGFD, which functionally mimic bona fide COL1A1-PDGFB fusions, leading presumably to a similar autocrine loop-stimulating PDGFRB. This study also emphasizes that COL1A1-PDGFBfusions can be cytogenetically cryptic on FISH testing in a subset of cases, thereby representing a diagnostic pitfall that pathologists should be aware of.

Secretory breast carcinoma (SBC) is a rare breast carcinoma with distinctive morphologic features and a recurrent specific chromosomal translocation t(12;15)(p13;q25), usually of low histologic grade and favorable prognosis. We describe the morphologic and genetic characteristics of 11 cases of SBC from 10 patients. Histologic and immunohistochemical analyses, fluorescence in situ hybridization using break-apart probes specific to ETV6on 12p13, reverse transcription polymerase chain reaction with in-house probes specific to the ETV6-NTRK3gene fusion, and DNA copy number variation by array comparative genomic hybridization analyses were performed on all cases. Seven cases were of low histologic grade, 3 were intermediate, and 1 had high-grade nuclear atypia, necrosis, and numerous mitoses. This patient had a fatal outcome. Five cases displayed low hormonal receptor expression, whereas the rest had basal-type immunoprofiles. All interpretable cases harbored an ETV6-NTRK3gene fusion by reverse transcription polymerase chain reaction and/or an ETV6rearrangement by fluorescence in situ hybridization, with duplication of the oncogenic derivative in 2 cases. Array comparative genomic hybridization analysis showed simplex genomic profiles. The 2 cases with ETV6-NTRK3duplication included a gain of 12p starting from the ETV6locus to the telomere, associated with a gain of the 15q from the centromere to NTRK3in 1 case, and in the other a normal profile up to NTRK3on 15q, and then a loss up to the telomere, suggesting loss of corresponding normal chromosome 15. These findings provide a novel insight into the morphologic and genetic spectrum of SBC, ranging from low-grade to high-grade histology, with occasional low hormonal receptor expression, simplex genomic profiles, and possible unfavorable course.

Ovarian small cell carcinoma of the hypercalcemic type (SCCOHT)/ovarian rhabdoid tumor is a rare and highly malignant tumor that typically occurs in young women. Up until now the diagnosis has been made on the basis of morphology without any specific immunohistochemical (IHC) markers. However, several authors have shown recently that SCCOHTs are characterized by inactivation of the SMARCA4gene (encoding the BRG1 protein) resulting in a loss of BRG1 protein expression in IHC. We evaluated BRG1 and INI1 expression in 12 SCCOHTs and in a series of 122 tumors that could mimic SCCOHT morphologically: 9 juvenile granulosa cell tumors, 47 adult granulosa cell tumors, 33 high-grade ovarian serous carcinomas, 9 desmoplastic round cell tumors, 13 Ewing sarcomas (5 from the pelvis and 8 from soft tissues), 1 round cell sarcoma associated with CIC-DUX4translocation from soft tissue (thigh), 1 case of high-grade endometrial stromal sarcoma of the ovary, and 9 melanomas. Forty-four adult granulosa cell tumors were interpretable by IHC. All 12 SCCOHTs were devoid of BRG1 expression and expressed INI1. All other interpretable 119 tumors showed BRG1 nuclear positivity, with variable staining proportions, ranging from 10% to 100% of positive cells (mean: 77%, median: 80%), variable intensities (weak: 5%, moderate: 37%, strong: 58%), and distributions: diffuse in 82 cases (70%) and heterogenous in 36 cases (30%). BRG1 positivity was heterogenous in desmoplastic round cell tumors and adult granulosa cell tumors. Overall, BRG1 is a useful diagnostic marker in SCCOHT, showing the absence of expression in SCCOHT. Nevertheless, the possible heterogeneity and the variable intensity of this staining warrant caution in the interpretation of BRG1 staining in biopsy specimens.

Neoadjuvant systemic therapy is being used increasingly in the treatment of early-stage breast cancer. Response, in the form of pathological complete response, is a validated and evaluable surrogate end point of survival after neoadjuvant therapy. Thus, pathological complete response has become a primary end point for clinical trials. However, there is a current lack of uniformity in the definition of pathological complete response. A review of standard operating procedures used by 28 major neoadjuvant breast cancer trials and/or 25 sites involved in such trials identified marked variability in specimen handling and histologic reporting. An international working group was convened to develop practical recommendations for the pathologic assessment of residual disease in neoadjuvant clinical trials of breast cancer and information expected from pathology reports. Systematic sampling of areas identified by informed mapping of the specimen and close correlation with radiological findings is preferable to overly exhaustive sampling, and permits taking tissue samples for translational research. Controversial areas are discussed, including measurement of lesion size, reporting of lymphovascular space invasion and the presence of isolated tumor cells in lymph nodes after neoadjuvant therapy, and retesting of markers after treatment. If there has been a pathological complete response, this must be clearly stated, and the presence/absence of residual ductal carcinoma in situmust be described. When there is residual invasive carcinoma, a comment must be made as to the presence/absence of chemotherapy effect in the breast and lymph nodes. The Residual Cancer Burden is the preferred method for quantifying residual disease in neoadjuvant clinical trials in breast cancer; other methods can be included per trial protocols and regional preference. Posttreatment tumor staging using the Tumor–Node–Metastasis system should be included. These recommendations for standardized pathological evaluation and reporting of neoadjuvant breast cancer specimens should improve prognostication for individual patients and allow comparison of treatment outcomes within and across clinical trials.

Neoadjuvant systemic therapy is being used increasingly in the treatment of early-stage breast cancer. Response, in the form of pathological complete response, is a validated and evaluable surrogate end point of survival after neoadjuvant therapy. Thus, pathological complete response has become a primary end point for clinical trials. However, there is a current lack of uniformity in the definition of pathological complete response. A review of standard operating procedures used by 28 major neoadjuvant breast cancer trials and/or 25 sites involved in such trials identified marked variability in specimen handling and histologic reporting. An international working group was convened to develop practical recommendations for the pathologic assessment of residual disease in neoadjuvant clinical trials of breast cancer and information expected from pathology reports. Systematic sampling of areas identified by informed mapping of the specimen and close correlation with radiological findings is preferable to overly exhaustive sampling, and permits taking tissue samples for translational research. Controversial areas are discussed, including measurement of lesion size, reporting of lymphovascular space invasion and the presence of isolated tumor cells in lymph nodes after neoadjuvant therapy, and retesting of markers after treatment. If there has been a pathological complete response, this must be clearly stated, and the presence/absence of residual ductal carcinoma in situ must be described. When there is residual invasive carcinoma, a comment must be made as to the presence/absence of chemotherapy effect in the breast and lymph nodes. The Residual Cancer Burden is the preferred method for quantifying residual disease in neoadjuvant clinical trials in breast cancer; other methods can be included per trial protocols and regional preference. Posttreatment tumor staging using the Tumor–Node–Metastasis system should be included. These recommendations for standardized pathological evaluation and reporting of neoadjuvant breast cancer specimens should improve prognostication for individual patients and allow comparison of treatment outcomes within and across clinical trials.

Le diagnostic de lésions prolifératives bénignes telles la cicatrice radiaire et le papillome, et de lésions prolifératives avec atypies ont augmenté ces dernières années avec la multiplication du nombre de prélèvements percutanés. De nombreux progrès ont été faits dans leur diagnostic histologique, leur classification et leur pathogenèse. Mais il n’existe toujours pas de consensus sur leur prise en charge, en particulier chirurgicale, qui reste controversée. Il est essentiel que radiologues et pathologistes communiquent pour définir ensemble la conduite à tenir.

De nouvelles recommandations internationales pour l’évaluation du statut HER2 dans les cancers du sein, basées sur plus de dix ans d’expérience et sur les résultats d’études cliniques et de concordance entre les différentes techniques de détection, viennent tout juste de voir le jour. Le présent article a pour objet de faire le point sur ces nouvelles recommandations, à la lumière de la publication récente du groupe de travail de l’American Society of Clinical Oncology (ASCO) et du Collège des pathologistes américains (CAP), adaptées à la pratique de la pathologie en France et revues par le groupe GEFPICS. À l’ère de la médecine personnalisée, la détermination du statut HER2 reste un élément phare dans le panel des biomarqueurs théranostiques des cancers du sein. Si l’interprétation du statut HER2 dans les cancers du sein est aisée dans la majorité des cas, un certain nombre de situations anatomocliniques est d’interprétation plus délicate, telles que la possibilité rare mais réelle de l’hétérogénéité intra-tumorale du statut de HER2, les formes à différenciation micropapillaire ou la ré-évaluation du statut des biomarqueurs lors de la rechute métastatique. Ces nouvelles recommandations abordent ces différentes questions, reprécisent les conditions pré-analytiques optimales et les critères d’interprétation (notamment des cas 2+), afin de réduire au maximum le risque de faux négatifs. Plus que jamais, la mobilisation de la spécialité d’anatomo-cytopathologie autour de la qualité des tests théranostiques témoigne de son implication dans la chaîne des soins en cancérologie.

Les tumeurs fixées et incluses en paraffine sont quotidiennement utilisées pour l’évaluation des biomarqueurs nécessaires au traitement des patientes atteintes d’un cancer du sein invasif. Les nouvelles recommandations internationales sur la phase pré-analytique ont été récemment revues, confirmant l’importance de la prise en charge optimale des prélèvements pour garantir des tests d’immunohistochimie ou d’hybridation in situ de qualité, quel que soit le biomarqueur envisagé. Incluant les procédés de fixation et de préparation des tissus, toutes les procédures pré-analytiques doivent être validées, standardisées et tracées. Elles nécessitent la collaboration et la formation de toutes les personnes impliquées dans le circuit du prélèvement, du préleveur jusqu’au technicien de pathologie et au pathologiste en passant par l’infirmière, ou le coursier. La prise en charge initiale optimale des pièces et une fixation de qualité sont des étapes majeures à maîtriser dans la phase pré-analytique. Cette mise à jour des recommandations du groupe d’étude des facteurs pronostiques immunohistochimiques dans le cancer du sein (GEFPICS) détaille et commente les différentes étapes pré-analytiques. L’observation de ces règles de bonne pratique, l’utilisation rigoureuse de témoins internes et externes et la participation régulière à des programmes d’assurance qualité sont autant de garanties pour une évaluation correcte et pérenne des biomarqueurs oncothéranostiques.

Endometrial stromal sarcomas represent the second most common mesenchymal uterine tumor. The 2003 WHO classification distinguishes low-grade and undifferentiated endometrial stromal sarcomas with different prognoses. Endometrial stromal sarcomas are a genetically heterogeneous group of sarcomas harboring different cytogenetic anomalies. Recently, a fusion between the YWHAEand FAM22A/Bgenes subsequent to a t(10;17) (q22;p13) has been described in endometrial sarcomas with high-grade histology. We examined YWHAErearrangements by FISH break-apart and RT-PCR in a series of 27 undifferentiated uterine stromal sarcoma without JAZF1rearrangements. Immunohistochemistry (IHC) was carried out with a panel of antibodies (estrogen (ER) and progesterone (PR) receptors, CD10, Cyclin D1, β-catenin, p53, and Ki-67). We identified a subgroup of endometrial sarcomas with high-grade histology and uniform morphology harboring YWHAErearrangements. FISH break-apart was interpretable in 20 cases (74%). Twelve cases (60%) showed <10% of tumor cells with a YWHAErearrangement, 4 cases (20%) showed between 10 and ≤20%, and 4 (20%) >20%. RT-PCR was tested on 24/27 cases (88%) and 19 cases were interpretable (79%). Five cases (26%) showed a specific fusion transcript YWHAE–FAM22A/Bsequence. The best concordance rate between FISH and RT-PCR (94%) was obtained with the threshold of 20% of cells with a YWHAErearrangement. The YWHAE-rearranged cases showed high-grade morphology with uniform appearance, spindle or round epithelioid cells, low ER and PR, CD10 expression, and a high and diffuse positivity for Cyclin D1, p53, and nuclear β-catenin negativity. Cyclin D1 was the most sensitive marker for high-grade endometrial sarcomas with YWHAErearrangement. All undifferentiated uterine sarcomas with pleomorphic appearances did not harbor any YWHAErearrangements, except for one case. Overall, for endometrial sarcoma cases with high-grade morphology we recommend to test for YWHAErearrangements by FISH break-apart, a cost- and time-efficient method, and to complete the investigation by RT-PCR in borderline cases.

Endometrial stromal sarcomas represent the second most common mesenchymal uterine tumor. The 2003 WHO classification distinguishes low-grade and undifferentiated endometrial stromal sarcomas with different prognoses. Endometrial stromal sarcomas are a genetically heterogeneous group of sarcomas harboring different cytogenetic anomalies. Recently, a fusion between the YWHAE and FAM22A/B genes subsequent to a t(10;17) (q22;p13) has been described in endometrial sarcomas with high-grade histology. We examined YWHAE rearrangements by FISH break-apart and RT-PCR in a series of 27 undifferentiated uterine stromal sarcoma without JAZF1 rearrangements. Immunohistochemistry (IHC) was carried out with a panel of antibodies (estrogen (ER) and progesterone (PR) receptors, CD10, Cyclin D1, β-catenin, p53, and Ki-67). We identified a subgroup of endometrial sarcomas with high-grade histology and uniform morphology harboring YWHAE rearrangements. FISH break-apart was interpretable in 20 cases (74%). Twelve cases (60%) showed <10% of tumor cells with a YWHAE rearrangement, 4 cases (20%) showed between 10 and ≤20%, and 4 (20%) >20%. RT-PCR was tested on 24/27 cases (88%) and 19 cases were interpretable (79%). Five cases (26%) showed a specific fusion transcript YWHAE–FAM22A/B sequence. The best concordance rate between FISH and RT-PCR (94%) was obtained with the threshold of 20% of cells with a YWHAE rearrangement. The YWHAE-rearranged cases showed high-grade morphology with uniform appearance, spindle or round epithelioid cells, low ER and PR, CD10 expression, and a high and diffuse positivity for Cyclin D1, p53, and nuclear β-catenin negativity. Cyclin D1 was the most sensitive marker for high-grade endometrial sarcomas with YWHAE rearrangement. All undifferentiated uterine sarcomas with pleomorphic appearances did not harbor any YWHAE rearrangements, except for one case. Overall, for endometrial sarcoma cases with high-grade morphology we recommend to test for YWHAE rearrangements by FISH break-apart, a cost- and time-efficient method, and to complete the investigation by RT-PCR in borderline cases.

Preoperative breast cancer diagnosis on core biopsies has become a standard of care in many countries. Controversies exist concerning the accuracy of HER2 testing on biopsies as compared with surgical specimens, and few data exist concerning the use of emerging technologies such as bright-field in-situhybridization in such a setting. A French multicenter, cross-sectional, histopathological study assessed the concordance of HER2 status determined by immunohistochemistry and silver (SISH) or chromogenic in-situhybridization (CISH) on core-needle biopsies with HER2 status determined by fluorescence in-situhybridization (FISH) on surgical specimens. The concordance between biopsy and operative results was also assessed for each method. We studied 260 breast tumors from 24 centers between April 2003 and August 2009. Excellent concordance (κ: 0.92–0.97) was shown between immunohistochemistry and FISH with low discordance rates (2–4%), high specificity (97–98%) and sensitivity values (95–99%), with no significant difference according to the immunohistochemistry interpretation guidelines used. The correlation between SISH and CISH on biopsies and FISH on surgical samples was strong (κ: 0.96 and 0.94, respectively), with no significant difference between false negative rates or sensitivity and specificity values (2 and 5%, 99 and 96%, 98 and 98%, respectively). Whatever the evaluation technique, excellent concordance between biopsies and surgical specimens was observed (κ⩾0.97; discordance rates between 1 and 2%), with high sensitivity (98–99%) and specificity (98–100%). Based on these results, when FISH cannot be used, SISH and/or CISH could be proposed as an alternative method to determine HER2 status and to confirm any ambiguous immunohistochemistry results, either for preoperative percutaneous biopsies or for surgical specimens. They could also be used for quality controls and immunohistochemistry calibration.

Preoperative breast cancer diagnosis on core biopsies has become a standard of care in many countries. Controversies exist concerning the accuracy of HER2 testing on biopsies as compared with surgical specimens, and few data exist concerning the use of emerging technologies such as bright-field in-situ hybridization in such a setting. A French multicenter, cross-sectional, histopathological study assessed the concordance of HER2 status determined by immunohistochemistry and silver (SISH) or chromogenic in-situ hybridization (CISH) on core-needle biopsies with HER2 status determined by fluorescence in-situ hybridization (FISH) on surgical specimens. The concordance between biopsy and operative results was also assessed for each method. We studied 260 breast tumors from 24 centers between April 2003 and August 2009. Excellent concordance (κ: 0.92–0.97) was shown between immunohistochemistry and FISH with low discordance rates (2–4%), high specificity (97–98%) and sensitivity values (95–99%), with no significant difference according to the immunohistochemistry interpretation guidelines used. The correlation between SISH and CISH on biopsies and FISH on surgical samples was strong (κ: 0.96 and 0.94, respectively), with no significant difference between false negative rates or sensitivity and specificity values (2 and 5%, 99 and 96%, 98 and 98%, respectively). Whatever the evaluation technique, excellent concordance between biopsies and surgical specimens was observed (κ 0.97; discordance rates between 1 and 2%), with high sensitivity (98–99%) and specificity (98–100%). Based on these results, when FISH cannot be used, SISH and/or CISH could be proposed as an alternative method to determine HER2 status and to confirm any ambiguous immunohistochemistry results, either for preoperative percutaneous biopsies or for surgical specimens. They could also be used for quality controls and immunohistochemistry calibration.

The necessity of excision is debatable when atypia are diagnosed at stereotactic vacuum-assisted breast biopsy (microbiopsy). Among the 287 surgical excisions performed at Institut Bergonié from 1999 to 2009, we selected a case–control study group of 151 excisions; 52 involving all the diagnosed cancers and 99 randomly selected among the 235 excisions without cancer, following atypical microbiopsy (24 flat epithelial atypia; 50 atypical ductal hyperplasia; 14 lobular neoplasia; 63 mixed lesions). Mammographical calcification (type, extension, complete removal) and histological criteria of epithelial atypia (type, number of foci, size/extension), topography and microcalcification extension at microbiopsy were compared according to the presence or absence of cancer at excision. Factors associated with cancer at excision were Breast Imaging Reporting and Data System (BI-RADS5) lesions, large and/or multiple foci of mammographical calcifications, histological type, number, size and extension of atypical foci. Flat epithelial atypia alone was never associated with cancer at excision. BI-RADS5, atypical ductal hyperplasia (alone or predominant) and >3 foci of atypia were identified as independent pejorative factors. There was never any cancer at excision when these pejorative factors were absent (n=31). Presence of one (n=59), two (n=23) or three (n=14) factors was associated with cancer in 24, 15 and 13 cases with an odds ratio=5.8 (95% CI: 3–11.2) for each additional factor. We recommend that mammographical data and histological characteristics be taken into account in the decision-making process after diagnosis of atypia on microbiopsy. With experienced senologists and strict histological criteria, some patients could be spared surgery resulting in significant patient, financial and time advantages.

The necessity of excision is debatable when atypia are diagnosed at stereotactic vacuum-assisted breast biopsy (microbiopsy). Among the 287 surgical excisions performed at Institut Bergonié from 1999 to 2009, we selected a case–control study group of 151 excisions; 52 involving all the diagnosed cancers and 99 randomly selected among the 235 excisions without cancer, following atypical microbiopsy (24 flat epithelial atypia; 50 atypical ductal hyperplasia; 14 lobular neoplasia; 63 mixed lesions). Mammographical calcification (type, extension, complete removal) and histological criteria of epithelial atypia (type, number of foci, size/extension), topography and microcalcification extension at microbiopsy were compared according to the presence or absence of cancer at excision. Factors associated with cancer at excision were Breast Imaging Reporting and Data System (BI-RADS5) lesions, large and/or multiple foci of mammographical calcifications, histological type, number, size and extension of atypical foci. Flat epithelial atypia alone was never associated with cancer at excision. BI-RADS5, atypical ductal hyperplasia (alone or predominant) and >3 foci of atypia were identified as independent pejorative factors. There was never any cancer at excision when these pejorative factors were absent (n=31). Presence of one (n=59), two (n=23) or three (n=14) factors was associated with cancer in 24, 15 and 13 cases with an odds ratio=5.8 (95% CI: 3–11.2) for each additional factor. We recommend that mammographical data and histological characteristics be taken into account in the decision-making process after diagnosis of atypia on microbiopsy. With experienced senologists and strict histological criteria, some patients could be spared surgery resulting in significant patient, financial and time advantages.

Pathologic diagnosis requires tissue fixation for histologic and immunohistologic analysis, and formalin is routinely used for this. The disadvantage of this fixative is its inability to preserve nucleic acids. Pathologic tumor diagnosis requires extensive molecular analyses, for which formalin fixation may be not adequate. Recently, an alcohol-based fixative (molecular fixative, MF) was described that allows nucleic acid preservation as well as histologic and immunohistologic studies. Moreover, the MF fixation processing system (Xpress) is fast and is well adapted to a routine process. We evaluated RNA and DNA quality within 1 month and after 1 year for 10 breast carcinomas and 20 sarcomas fixed in MF in comparison with the corresponding frozen tumors. The quality of DNA extracted from the MF-fixed tissue was similar to that extracted from the frozen tumors. The quality of RNA extracted from the MF-fixed tissue was lower than that of frozen tumors; nevertheless, a majority of RNA integrity number (RIN) values were greater than 7. Gene expression quantification by real-time polymerase chain reaction gave comparable results between tumors fixed with MF and frozen tumors. Tissue fixation at 4°C with the MF improved the RNA quality measured by the RIN value. However, after storage for 1 year at room temperature, although DNA quality was preserved, RNA extracted from tissues fixed with the MF was degraded. Tissue fixation with the MF is an important improvement for molecular pathologic diagnosis, enabling a combination of routine pathologic diagnoses and current molecular diagnoses if they are carried out near the processing time.

L’analyse extemporanée des ganglions sentinelles (GAS), prélevés dans les carcinomes infiltrants du sein, permet, en cas d’envahissement métastatique, de réaliser un curage axillaire dans le même temps opératoire. Or, le principe, l’efficacité et la méthode d’analyse extemporanée (cytologique, histologique) sont controversés. Le but de notre étude a été d’évaluer la pertinence de l’examen extemporané cytologique pratiqué dans notre institut.

Breast carcinoma is the main malignant tumor occurring in patients with Cowden disease, a cancer-prone syndrome caused by germline mutation of the tumor suppressor gene PTENcharacterized by the occurrence throughout life of hyperplastic, hamartomatous and malignant growths affecting various organs. The absence of known histological features for breast cancer arising in a PTEN-mutant background prompted us to explore them for potential new markers. We first performed a microarray study of three tumors from patients with Cowden disease in the context of a transcriptomic study of 74 familial breast cancers. A subsequent histological and immunohistochemical study including 12 additional cases of Cowden disease breast carcinomas was performed to confirm the microarray data. Unsupervised clustering of the 74 familial tumors followed the intrinsic gene classification of breast cancer except for a group of five tumors that included the three Cowden tumors. The gene expression profile of the Cowden tumors shows considerable overlap with that of a breast cancer subgroup known as molecular apocrine breast carcinoma, which is suspected to have increased androgenic signaling and shows frequent ERBB2amplification in sporadic tumors. The histological and immunohistochemical study showed that several cases had apocrine histological features and expressed GGT1, which is a potential new marker for apocrine breast carcinoma. These data suggest that activation of the ERBB2-PI3K-AKT pathway by loss of PTEN at early stages of tumorigenesis promotes the formation of breast tumors with apocrine features.

Most Her2 testing guidelines recommend that all cases scoring Her2 2+ by immunohistochemistry should be analyzed by fluorescent in situhybridization (FISH) to determine HER2status to confirm eligibility for Trastuzumab therapy in breast cancer. The aim of our study was to determine HER2gene and chromosome 17 (CEN17) status in a series of 108 Her2 2+ consecutive cases and study the correlation between pathological characteristics of the tumors and HER2 amplification. Invasive breast cancers were tested by FISH using the Dako HER2FISH pharmDx® kit. The Her2 immunohistochemistry protocol was performed using the polyclonal AO485 antibody (Dako®) diluted to 1:1500. HER2and CEN17 status were correlated to tumor SBR grade, mitotic count, estrogen receptor, progesterone receptor status and percentage of Her2 immunohistochemistry-positive cells. Following Food and Drug Administration guidelines, ie, HER2/CEN17 ratio ≥2 and an HER2copy number >4, amplified cases were observed in 36 (33%) and 49 (45%) cases, respectively, and following American Society of Clinical Oncology/College of American Pathologists guidelines, ie, HER2/CEN17 ratio >2.2 and an HER2copy number >6, amplified cases represented 30 and 24% of the study population, respectively. Chromosome 17 polysomy (CEN17 >2.25) was observed in 39 (36%) tumors. Significant positive correlations were found between FISH HER2amplified cases and Her2 immunostaining >60% (P=1.1.10−5), SBR grade 3 (P=0.0001), nuclear atypia (P=0.03) and mitotic count (P=0.008). By multivariate analysis, Her2 immunostaining >60% (P<10−3) and SBR grade 3 (P<10−3) were independent factors predicting HER2amplification status irrespective to cutoff guidelines. All SBR grade 3 cases with more than 60% Her2+ cells had an HER2/CEN17 ratio ≥2, only one had a ratio ≤2.2. In our series of consecutive Her2 2+ cases, one-third demonstrated HER2amplification, and one-third had chromosome 17 polysomy. Pathological factors, in particular SBR grade 3 and more than 60% Her2+ cells, were significantly correlated with HER2amplification.

Peritumoral emboli assessed on hematoxylin–eosin-stained slides are taken into account for treatment of patients with operable breast cancer. We assessed whether immunostaining with D2-40 improves the prognostic significance of emboli in a group of tumors with a large immunohistochemical sampling and a long-term follow-up. Topography, number, and extension of hematoxylin–eosin and D2-40 emboli were compared in 94 node-negative breast cancers (median number of immunostained slides per tumor: 3). Metastatasis-free survival of patients with or without hematoxylin–eosin and/or D2-40 emboli were evaluated (median follow-up of 178 months). Hematoxylin–eosin emboli were detected in 14 (15%) tumors and were located at distance from the tumor. D2-40 emboli were detected in 39 (41%) tumors and was often multiple (n=30), extensive (n=23), located within (n=13), close to (n=10) or at distance from the tumor (n=16). The 12 distant hematoxylin–eosin and D2-40 emboli were located in the same vessels (seven missed at the first hematoxylin–eosin examination and secondarily diagnosed by D2-40 staining). A difference in metastasis-free survival was found only between patients with no D2-40 emboli and those with distant D2-40 emboli (P=0.02). D2-40 emboli located within or close to the tumor had no prognostic value. Comparing the metastasis-free survival of patients with or without hematoxylin–eosin emboli, the prognostically unfavorable significance of hematoxylin–eosin emboli was improved when taking into account the seven patients with missed emboli at the first examination and secondarily diagnosed by D2-40 staining (P=0.006 vs0.003). To conclude, D2-40 increases the diagnostic sensitivity of emboli in breast carcinoma and the high incidence of D2-40 emboli might be related to the number of immunostained slides per case. Nevertheless, only distant D2-40+ emboli had a prognostic impact. In practice, D2-40 might be useful to detect missed hematoxylin–eosin emboli especially in cases without any other prognostically unfavorable criterion.

Typical medullary breast carcinoma (MBC) has recently been recognized to be part of the basal-like carcinoma spectrum, a feature in agreement with the high rate of TP53mutations previously reported in MBCs. The present study was therefore designed to identify phenotypic and genetic alterations that distinguish MBCs from basal-like carcinomas (BLC). Expression levels of estrogen receptor (ER), progesterone receptor (PR), ERBB2, TP53, cytokeratins (KRTs) 5/6, 14, 8/18, epidermal growth factor receptor and KIT, as well as TP53gene sequence and high-density array comparative genomic hybridization (CGH) profiles, were assessed and compared in a series of 33 MBCs and 26 BLCs. All tumors were negative for ER, PR and ERBB2. KRTs 5/6 were more frequently expressed in MBCs (94%) than in BLCs (56%) (p= 0.0004). TP53mutations were disclosed in 20/26 MBCs (77%) and 20/24 BLCs (83%). Array CGH analysis showed that a higher number of gains (95 regions) and losses (34 regions) was observed in MBCs than in BLCs (36 regions of gain; 13 regions of losses). In addition, gains of 1q and 8q, and losses of X were found to be common to the two groups, whereas gains of 10p (53% of the cases), 9p (30.8% of the cases) and 16q (25.8% of the cases), and losses of 4p (34.8% of the cases), and amplicons of 1q, 8p, 10p and 12p were the genetic alterations found to characterize MBC. Our study has revealed that MBCs are part of the basal-like group and share common genomic alterations with BLCs, the most frequent being 1q and 8q gains and X losses; however, MBCs are a distinct entity within the basal-like spectrum, characterized by a higher rate of KRT 5/6 expression, a higher rate of gains and losses than BLCs, recurrent 10p, 9p and 16q gains, 4p losses, and 1q, 8p, 10p and 12p amplicons. Our results thus contribute to a better understanding of the heterogeneity in basal-like breast tumors and provide potential diagnostic tools.

The significance of microinvasion is still debated and clinical management is controversial. The authors defined ductal carcinoma in situ with microinvasion (DCIS-MI) as DCIS with infiltration of the periductal stroma by a few tumor cells, singly (type 1) or in clusters (type 2). With this definition, the authors attempted to evaluate the clinical significance of microinvasion. The authors compared the clinical, pathologic features, and survival (median follow-up, 7.3 years) of 1248 patients with, respectively, DCIS (722 patients), DCIS-MI with microinvasion type 1 and type 2 (243 patients), and invasive ductal carcinoma in situ with a predominant DCIS component greater than or equal to 80% of the tumor (IDC-DCIS, 283 patients). Microinvasion was associated with DCIS histologic type, grade, and extent (respectively, P &lt; 10⁻⁸, P &lt; 10⁻³, P &lt; 10⁻⁴). Axillary lymph node metastases were observed in a few patients with DCIS and DCIS-MI type 1 (respectively, 1.4% and none), in 10.1% with DCIS-MI type 2 and in 27.6% with IDC-DCIS. Metastasis free and overall survival probabilities were significantly different between three groups in the following order from best to worst prognosis: 1) the group comprising DCIS and DCIS-MI type 1, 2) the DCIS-MI type 2 group, and 3) the IDC-DCIS group. The authors' results suggest there are two types of DCIS-MI: 1) type 1 that behaves like DCIS and should be managed as such; 2) type 2 that is less pejorative than IDC-DCIS but is more so than type 1. Cancer 2002;94:2134–42. © 2002 American Cancer Society. DOI 10.1002/cncr.10451

To evaluate the prognostic relevance of Ki‐67 and topoisomerase IIα expression in relation to tumor stage, grade, and hormone receptor content, 942 ductal infiltrating carcinomas of the breast were examined by means of the monoclonal antibodies Ki‐S11 (Ki‐67) and Ki‐S4 (topoisomerase IIα). pS2, c‐erbB2, and p53 were additionally considered as prognostic variables. The median follow‐up time was 149 months. Eight‐hundred‐and‐sixty‐three tumors reacted with Ki‐S11 and Ki‐S the labeling indices of the two antigens were closely associated (r=0.93). Both correlated positively with the tumor size, c‐erbB2, and p53 expression, and negatively with patient age, hormone receptor content, and pS2 immunostaining. In the univariate analysis, Ki‐S11 and Ki‐S4 scores, nodal status, tumor size, tumor grade, and progesterone receptor content strongly predicted both overall and metastasis‐free survival (p <0.00001). Estrogen receptor status, p53, and c‐erbB2 were of minor significance. Concerning overall survival, multivariate Cox regression analysis selected a Ki‐S4 score >25% (p < 0.00001) next to the nodal status, and before tumor size, progesterone receptor content, and patient age. Independent predictors of the occurrence of distant metastases were nodal status, Ki‐S4, tumor size, grade 1, and progesterone receptor negativity, in that order. The Ki‐S11 score was of independent prognostic significance only if examined as a continuous variable. We conclude that topoisomerase IIα expression as assessed by monoclonal antibody Ki‐S4 may add valuable information to current prognostic models for breast cancer. Its predictive value appears to be essentially related to the proliferative activity of tumor cells.

P53 immunohistochemical detection using DO7 antibody on 942 cases of previously untreated breast invasive ductal carcinoma (IDC) with a median follow up of 117.9 months (89 to 160) was performed.

We developed a scanning laser terahertz (THz) near-field reflection imaging system using a fiber-coupled femtosecond laser source for high spatial resolution measurements without any sub-wavelength probes. In this system, THz wave pulses generated at the laser focusing spot in a non-linear optical crystal interact with a sample set on the crystal. Using this system, we obtained the THz reflection images of metallic structures with a resolution of ??THz/34 (?THz= 680 ?m) and also observed paraffin-embedded human breast tissue of several 10 ?m in size. These results promise that this system can be suitable for industrial applications which require micron-scale resolution.