Bone metastasis is strongly associated with estrogen receptor-positive/progesterone receptor-negative breast carcinomas

Bone is one of the most common sites of distant metastasis for breast carcinomas. In this study, our objective is to identify molecular markers and molecular subtypes that may predict patients at higher risk of developing bone metastasis. Immunohistochemical analysis with antibodies against estrogen receptor α, progesterone receptor, androgen receptor, Her2/neu, epidermal growth factor receptor, CK5/6, CK14, CK17, CK8, and CK18 was performed on representative sections of 21 breast carcinomas with bone metastasis and 94 cases without bone metastasis. The expression rates of receptors, subtype distributions (basal versus nonbasal) of 3 molecular classifications (cytokeratin, triple negative, and cytokeratin/triple negative), and 5 subtypes of cytokeratin/triple negative classification were compared between these 2 groups. We found that (1) the breast cancers with bone metastasis were associated with a significant percentage of estrogen receptor–positive/progesterone receptor–negative tumors compared with tumors without bone metastasis (38% versus 6%, P < .0001). (2) There was significant difference on estrogen receptor expression between high grade and non–high grade in tumors with or without bone metastasis (P = .0084 and 1.0000, respectively). (3) The breast cancers with bone metastasis were more likely to be estrogen receptor positive (85%) and androgen receptor positive (95%) compared with those without bone metastasis (59% and 74%, respectively, both P < .05). (4) There was no significant difference between tumors with or without bone metastasis in subtype distribution (basal versus nonbasal) among all 3 molecular classifications. (5) Luminal B carcinomas of cytokeratin/triple negative classification tended to be associated with bone metastasis but not to a statistically significant extent. In conclusion, bone metastasis is strongly associated with estrogen receptor–positive/progesterone receptor–negative tumors. Significant difference in estrogen receptor expression between high-grade and non–high-grade tumors with bone metastasis suggests that different panels of molecular markers should be used to predict bone metastasis in these 2 groups of tumors.     

Peptidomic analysis of breast cancer reveals a putative surrogate marker for estrogen receptor-negative carcinomas

Estrogen-receptor status provides a major biomarker in breast cancer classification and has an important impact on prognosis and treatment options. The aim of this study was to investigate peptide profiles of invasive breast cancer with positive (n=39) and negative receptor status (n=41). Peptide profiles were generated by 'Differential Peptide Display', which is an offline-coupled combination of reversed-phase-HPLC and MALDI mass spectrometry. Mass spectrometric data were correlated with the immunohistochemically determined receptor state. Identification of peptides of interest was carried out by additional mass spectrometric methods (eg MALDI-TOF-TOF-MS-MS). Approximately 3000-7000 signals were detected per sample and thymosin alpha-1, an asparaginyl endopeptidase generated cleavage product of the ubiquitous acidic protein prothymosin-alpha, was found to differentiate the tumor samples according to their receptor status with the highest specificity. The concentration of Thymosin alpha-1 was found to be upregulated (n=37) in estrogen-negative cancer samples and downregulated (n=32) in estrogen-positive breast cancer samples. The expression of the precursor protein (Prothymosin-alpha) has been discussed previously as a prognostic factor in breast cancer. It is involved in the ER signal transduction pathway as an anti-coactivator-inhibitor. From our findings we conclude that Thymosin alpha-1 could serve as a surrogate marker in breast cancers and may indicate ER functionality.     

Pregnancy-associated plasma protein A and extensive necrosis. Clinically significant predictors of early recurrence in stage I estrogen receptor-negative breast carcinoma

Despite advances in the detection of early breast cancer, 25 to 35% of patients with stage I disease die from metastatic breast carcinoma. To identify those patients at risk for early recurrence, we reviewed 33 clinical and pathologic features as well as immunoperoxidase-staining characteristics for carcinoembryonic antigen, human chorionic gonadotrophin, pregnancy specific beta-1 glycoprotein, and pregnancy-associated plasma protein A (PAPP-A), in 40 patients with stage I estrogen receptor-negative breast carcinoma. Sixteen of the 40 patients (40%) developed tumor recurrence within 2 years. Pairwise correlations between recurrence and clinicopathologic features, including tumor marker immunostaining, revealed significant correlations between extensive necrosis, nuclear atypia, mitoses, and PAPP-A staining. In multivariant linear discriminant analysis, only PAPP-A staining and extensive necrosis entered as significant independent predictors. In the recurrent group, nine of 16 (56%) were PAPP-A positive compared with one of 24 (4%) in the nonrecurrent group (p less than 0.001), whereas nine of 16 (56%) contained extensive necrosis compared with three of 24 (11%) in the nonrecurrent group (p less than 0.005). When the independent risk factor of PAPP-A positivity and extensive necrosis were combined, 13 of 16 (81%) of the recurrent tumors were either PAPP-A positive or extensively necrotic compared with four of 24 (16%) of the nonrecurrent group. Thus, positive immunostaining for PAPP-A and the presence of extensive necrosis are clinically significant independent predictors of early recurrence in patients with stage I, estrogen receptor-negative breast carcinoma. These risk factors for early recurrence may be helpful in prospectively selecting patients most eligible to receive adjuvant chemotherapy.     

Estrogen receptor-negative,progesterone receptor-positive breast carcinoma: poor clinical outcome

OBJECTIVE: The biological significance of breast carcinomas negative (-) for estrogen receptor (ER), but positive (+) for progesterone receptor (PR) is unclear. It has been proposed that these tumors contain ER whose presence is masked in binding assays by endogenous estrogen. We analyzed the clinical outcome of 17 patients with ER-PR+ tumors. METHODS: The disease-free and overall survival of a series of 300 women with invasive breast carcinoma was followed for 7 to 79 (median 41) months. RESULTS: The recurrence rate was significantly greater in ER-PR+ tumors (8/17 [47%]) than in ER+PR+ tumors (27/177 [15%]), and it was similar to the high recurrence rate of ER-PR- tumors (21/57 [37%]). The cancer-related death rate was 3 1/2 times higher in the ER-PR+ group than in the ER+PR+ group. A significant association between ER-PR+ tumors and tumor recurrence or cancer-related death persisted even after correction for other variables associated with poor outcome (eg, tumor size and lymph node involvement). CONCLUSIONS: Estrogen receptor-negative, progesterone receptor-positive breast carcinomas are biologically different from ER+PR+ tumors and have a poor clinical outcome.     

Androgen receptor expression in estrogen receptor-negative breast cancer. Immunohistochemical, clinical, and prognostic associations

We sought to determine the prevalence of androgen receptor (AR) expression in a predominantly estrogen receptor (ER)-negative subset of breast cancers and delineate the immunohistochemical and clinical associations, including whether AR expression has prognostic significance in ER-negative tumors. We identified 69 ER-negative and 19 ER-positive breast cancer cases with concurrent immunohistochemical prognostic panels (ER, PR, HER-2/neu, Ki-67, and p53); immunohistochemical analysis was performed for AR using standard techniques. Clinical data were extracted from medical records. chi 2 tests were used to assess associations between variables. AR was found in 49% (34/69) of ER-negative and 89% (17/19) of ER-positive cases. In ER-negative tumors, AR was associated with increased age (P = .02), postmenopausal status (P < .001), tumor grade (P = .03), tumor size (P = .03), and HER-2/neu overexpression (P = .003). In ER-positive tumors, AR was associated with progesterone receptor expression (P < .03). In univariate analysis of ER-negative tumors, patients with AR-positive tumors had significantly better disease-free survival (P = .049). AR is expressed in a significant number of ER-negative cases and shows significant associations with important clinical and pathologic prognostic factors.     

乳腺大汗腺癌的形态学与免疫表型特征

目的 观察乳腺大汗腺癌(apocrine carcinoma，AC)的形态学和免疫表型特征，并探讨其诊断标准。方法 对HE染色片中显示有大汗腺细胞学特点的30例乳腺癌标本进行组织学观察和免疫组化S—P法检测，并选取5例大汗腺化生(apocrine metaplasia，AM)及2例大汗腺腺病(apocrine adenosis，AA)和AA伴导管原位癌(DCIS)1例标本作对照组。使用的-抗包括GCDFP—15、Mucin—1、AR、ER、C—erbB-2、Ki-67。结果 ①8例被确定为AC，均为高核级，其中大汗腺导管原位癌(ADCIS)1例、ADCIS伴浸润性大汗腺癌(IAC)5例，IAC伴导管原位癌(DCIS)2例。镜下AC细胞的共同特征为：细胞大，胞界清，胞质丰富，深嗜伊红染，颗粒状；核大、圆，核仁常大而突出。依据胞质形态可分为：A型，胞质深伊红染，有数量不等的清晰颗粒；B型，胞质内大量细空虚的小泡，可聚呈泡沫状，类似组织细胞或皮脂腺细胞；C型，胞质浅染、均质，如细磨砂玻璃样；②8例AC与5例AM及2例AA均共同表达GCDFP—15、AR、Mucin—1及Ki-67；除1例外，ER均不表达；③AC的GCDFP—15的表达与大汗腺化生不同：前者呈胞质内弥漫颗粒或空泡状，表达有异质性；后者阳性颗粒聚成小球状团簇，位于细胞质的核上区。④AR平均阳性细胞数在Ac及AM分别为33．4％及93．8%；Ki-67平均阳性细胞数在AC及AM分别为52．4％及40％。结论 AC的诊断须具备3项标准：①HE细胞形态成分90％以上符合Ac型细胞；②GCDFP—15阳性，阳性细胞数占癌细胞总数50％以上；⑧AR阳性，阳性细胞数大于癌细胞总数20％。     

乳腺导管内二态性乳头状癌的形态及免疫表型特征

目的探讨乳腺导管内二态性乳头状癌的形态及免疫表型特征。方法收集具有二态性形态学特征的乳腺导管原位癌7例,观察其组织形态学特点,并采用EnVision免疫组化法检测CK5/6、CK34βE12、CK8、CD10、p63、SMA、ER、E-cad、Syn、CgA、GCDFP-15和AR的表达。结果二态性导管内乳头状癌4例,具有二态性特征的筛状型导管原位癌3例;其中4例伴有导管内非典型增生。二态性乳头状癌有两种形态不同的肿瘤细胞:一种是普通性肿瘤细胞,多为柱状细胞,胞质不透明,核为低或中间级别。此类细胞分布于乳头或筛状结构的浅层;第二种细胞是二态性肿瘤细胞,多呈多角形或立方形,胞质透明或淡染,核级别及形态与普通性肿瘤细胞相同。此种细胞位于柱状肿瘤细胞内侧,排列呈条带状、筛状、实性巢状或单个细胞播散分布,其胞质及细胞分布位置类似增生的肌上皮细胞。两种肿瘤细胞均显示CK8、ER阳性,基底高分子量角蛋白(CK5/6、CK34βE12)、肌上皮(CD10、p63、SMA)、Syn和CgA阴性;E-cad阳性5例,另2例部分细胞呈胞膜胞质型微弱阳性而大部分细胞不表达;1例GCDFP-15及AR阳性。结论二态性导管内乳头状癌的两种肿瘤细胞均呈腺系上皮分化,两种肿瘤细胞的形态及胞质染色不同,但核的异型性及其形态相同,免疫组化结果也相同。二态性肿瘤细胞在HE切片内与肌上皮细胞很相似,易被误认为肌上皮,从而导致诊断不足或误诊,宜予高度重视。     

乳腺肌上皮癌3例临床病理特征分析

目的探讨乳腺肌上皮癌的临床表现、组织形态学改变和免疫表型特征。方法采用HE及免疫组化Leica BOND-MAX全自动染色仪Bond Polymer Refine Detection法,对3例乳腺肌上皮癌进行染色,并复习相关文献。结果乳腺肌上皮癌的癌组织多由梭形细胞组成,部分病例可见上皮样细胞、浆样细胞和透明细胞,呈实性片状、肺泡样、条索状排列;核分裂象易见;无坏死;可发生腋窝淋巴结转移。免疫表型:3例均表达CK、EMA、CD10、p40、actin、Calponin、p63;2例表达CK5/6、CK14、SMA;1例表达S-100蛋白、D2-40、CAM5.2;CD117、CD34、ER、PR、C-erb B-2/HER-2和desmin均不表达;Ki-67增殖指数为10%~20%。结论肌上皮癌细胞形态多样,结合免疫组化组合标记可做出明确诊断,治疗方式以乳腺癌根治术为主。     

Morphologic identity of primary tumor and axillary metastases in breast carcinoma. A quantitative study

The amount of glandular differentiation was quantitated in 64 cases of primary human mammary ductal carcinoma (PR) and their axillary lymph node metastases (LNMs) to identify differences between the two groups and their effect on patient prognosis. A significant difference between a PR and any of its LNMs was uncommon (less than 5% of the cases), and variation between each PR and its LNMs was directly proportional to the amount of differentiation within the PR itself. Variance among the LNMs was proportional to that found within the PR. Each LNM usually varied as much within itself as it did from the other LNMs. Prognosis was not affected by the LNMs being less differentiated than the PR, nor vice versa. This study has provided no evidence for selection of a morphologically more aggressive tumor clone during this phase of the metastatic process, but selection by other morphologic or functional parameters is not excluded.     

Demonstration of estrogen receptor by immunohistochemical staining in paraffin sections of breast carcinoma

Paraffin embedded sections of 64 breast carcinomas were stained immunohistochemically using a commercially available monoclonal antibody to estrogen receptor. To improve the sensitivity of the staining, the authors used a Pronase enzyme pretreatment, biotinylated antibody to rat IgG as secondary antibody, streptavidin-alkaline phosphatase as tertiary reagent and fast red as chromogen. When compared to the results of estrogen receptor enzyme immunoassay, this method yielded an 85.9% concordance rate, 86.2% specificity and 85.7% sensitivity. When compared to estrogen receptor immunocytochemistry(ER-ICA) in frozen section and considering the inherent advantages of immunohistochemical staining over biochemical assay, the major advantages of this method are good morphology, suitability for retrospective study and reduced cost of staining due to dilution of expensive primary antibody. Thus, this method offers an alternative to ER assay using fresh tissue and should provide additional valuable information about estrogen receptor.     

A methyl-deviator epigenotype of estrogen receptor-positive breast carcinoma is associated with malignant biology

We broadly profiled DNA methylation in breast cancers (n = 351) and benign parenchyma (n = 47) for correspondence with disease phenotype, using FFPE diagnostic surgical pathology specimens. Exploratory analysis revealed a distinctive primary invasive carcinoma subclass featuring extreme global methylation deviation. Subsequently, we tested the correlation between methylation remodeling pervasiveness and malignant biological features. A methyl deviation index (MDI) was calculated for each lesion relative to terminal ductal-lobular unit baseline, and group comparisons revealed that high-grade and short-survival estrogen receptor-positive (ER(+)) cancers manifest a significantly higher MDI than low-grade and long-survival ER(+) cancers. In contrast, ER(-) cancers display a significantly lower MDI, revealing a striking epigenomic distinction between cancer hormone receptor subtypes. Kaplan-Meier survival curves of MDI-based risk classes showed significant divergence between low- and high-risk groups. MDI showed superior prognostic performance to crude methylation levels, and MDI retained prognostic significance (P < 0.01) in Cox multivariate analysis, including clinical stage and pathological grade. Most MDI targets individually are significant markers of ER(+) cancer survival. Lymphoid and mesenchymal indexes were not substantially different between ER(+) and ER(-) groups and do not explain MDI dichotomy. However, the mesenchymal index was associated with ER(+) cancer survival, and a high lymphoid index was associated with medullary carcinoma. Finally, a comparison between metastases and primary tumors suggests methylation patterns are established early and maintained through disease progression for both ER(+) and ER(-) tumors.     

The evaluation of estrogen receptor in primary breast carcinoma by computer-assisted image analysis

A monoclonal antibody prepared against estrogen receptor has been shown to be specific and sensitive for the detection of estrogen receptor in human breast lesions by use of immunohistochemical methods. Two hundred selected cases of primary breast carcinoma were assayed for estrogen receptor content by biochemical and immunohistochemical procedures. Quantitative evaluation was by biochemical, immunohistochemical, and automated computer-assisted image analysis using the Cell Analysis System's CAS/100 machine (Lombard, IL). Quantitative estrogen receptor content was determined by dextran-coated charcoal analysis and sucrose density gradient analysis. Immunohistochemical evaluation incorporated both intensity and distribution of staining, yielding a subjective score, histologic score (HSCORE). An objective quantitation, also incorporating intensity and distribution of staining, was done by computer-assisted image analysis, quantitative immunocytochemical score (QIC SCORE). HSCORE analysis was done with and without methyl green counterstain with no loss of sensitivity. Comparison of QIC SCORE with the biochemical and immunohistochemical analysis of the tissues examined revealed excellent sensitivities and specificities. These data suggest that automated image analysis provides an effective qualitative and quantitative means of evaluating estrogen receptor content in human breast cancers.     

High-grade endometrial carcinomas: Morphologic spectrum and molecular classification

High-grade endometrial carcinoma (HGEC) is a heterogeneous group of tumors with various morphologic, genetic, and clinical characteristics. Morphologically, HGEC includes high-grade endometrioid carcinoma, serous carcinoma, clear cell carcinoma, undifferentiated/dedifferentiated carcinoma, and carcinosarcoma. The morphologic classification has been used for prognostication and treatment decisions. However, patient management based on morphologic classification is limited by suboptimal interobserver reproducibility, variable clinical outcomes observed within the same histotype, and frequent discordant histotyping/grading between biopsy and hysterectomy specimens. Recent studies from The Cancer Genome Atlas (TCGA) Research Network established four distinct molecular subtypes: POLE-ultramutated, microsatellite unstable, copy number high, and copy number low groups. Compared to histotyping, the TCGA molecular classification appears superior in risk stratification. The best prognosis is seen in the POLE-ultramutated group and the worst in copy number high group, while the prognosis in the microsatellite unstable and copy number low groups is in between. The TCGA subtyping is more reproducible and shows a better concordance between endometrial biopsy and resection specimens. It has now become apparent that the molecular classification can supplement histotyping in patient management. This article provides an overview of the pathologic diagnosis/differential diagnosis of HGEC and the TCGA classification of endometrial cancers, with the clinical significance and applications of TCGA classification briefly discussed when appropriate.     

Minimum formalin fixation time for consistent estrogen receptor immunohistochemical staining of invasive breast carcinoma

To identify the minimum time necessary for consistent immunohistochemical estrogen receptor (ER) results in our laboratory, we evaluated results in timed fixation blocks and cases with disparate and similar needle core biopsy and partial mastectomy specimens. Tissue sections of 24 ER-positive, invasive breast carcinomas were fixed for 3, 6, 8, and 12 hours and 1, 2, and 7 days. ER values were quantified using the Q score (0-7). In timed fixation blocks, the mean Q score per block was 2.46 for blocks fixed for 3 hours, 5.75 for blocks fixed for 6 hours, and 6.70 for blocks fixed for 8 hours (P < .001). The difference between the case maximum and mean block Q scores was a plateau of almost 0 at 6 to 8 hours of formalin fixation. For needle core biopsy specimen fixation times, the means for specimens with ER-disparate and ER-similar results were 1.2 and 6.3 hours, respectively (P = .01). The minimum formalin fixation time for reliable immunohistochemical ER results is 6 to 8 hours in our laboratory, regardless of the type or size of specimen.     

Estrogen receptor-negative breast carcinomas: a review of morphology and immunophenotypical analysis.

Estrogen receptor (ER)-negative breast cancers are a group of tumors with poor prognosis and fewer cancer prevention and treatment strategies compared to ER-positive tumors. The aim of this study was to assess the morphological characteristics and immunohistochemical profile of ER-negative tumors and thus to understand the biological behavior and unique nature. In total, 291 consecutive ER-negative cases available from our primary breast cancer series were examined. Hematoxylin- and eosin-stained sections of all the cases were studied for several morphological parameters and their immunophenotype profile. These findings were correlated with patient and tumor characteristics and survival data. ER-negative tumors constituted 30% of the primary operable breast cancer series. The majority of tumors were grade 3 (94%) and the commonest histological types were ductal/no specific type (85%), and atypical medullary carcinoma (8%). High-grade comedo-type necrosis, lymphoid stroma, central necrosis/fibrosis and pushing margins were the most common morphological features. The presence of a pushing margin showed a significant relation to androgen receptor negativity, absence of epidermal growth factor receptor expression and negative lymph nodes. Lymphoid stroma and comedo-necrosis correlated with higher tumor grade. ER-negative breast cancers are a distinct group of tumors with several common morphological features. Grade 3 histology, pushing margin, lymphoid stroma, comedo-type necrosis and central fibrosis/necrosis are the dominant morphological findings. The presence of a pushing margin appears to have a significant correlation with negative lymph node status. ER-negative tumors show a higher expression of p53, CerbB2 and epidermal growth factor receptor compared to ER-positive breast cancer. These unique features support the concept that ER-negative tumors are a morphologically and phenotypically distinct entity and provide a rationale for the study and use of newer promising agents in the treatment of ER-negative breast cancer.     

Image analysis for quantitation of estrogen receptor in formalin-fixed paraffin-embedded sections of breast carcinoma.

Monoclonal antibody to human estrogen receptor (ER) provides a useful immunohistochemical tool for the evaluation of ER content in breast carcinoma, but visual interpretation is subjective. Computer-assisted image analysis has proved effective in immunohistochemical quantitation of ER in fresh tumor imprints and cryostat sections. We examined the usefulness of this technique in 5-microns-thick formalin-fixed paraffin-embedded tissue sections of 66 cases of primary breast carcinoma previously assayed by dextran-coated charcoal (DCC) analysis. Immunohistochemistry was automated and performed on a Code-on slide stainer (Instrumentation Laboratories, Lexington, MA) using Pronase predigestion, a monoclonal antibody (ER-ICA; Abbott, Chicago, IL), and a biotin-labeled secondary antibody. Detection was achieved with an avidin-alkaline phosphatase conjugate and nitroblue tetrazolium (NBT) bromochloroindoyl phosphate (BCIP) substrate. The immunohistochemical ER staining was analyzed visually and with the CAS/200 image analyzer (Elmhurst, IL). The visual semiquantitative histologic scores (HSCORE), the automated quantitative assays including the percentage of positive nuclear areas (PNA), and the quantitative immunocytochemical scores (QIC SCORE = PNA x % of positive stain/10) were compared with the corresponding DCC results. Linear correlations were demonstrated between all immunohistochemical assays and the logarithm of DCC, the strongest correlation seen with PNA (r = 0.91). Threshold points for positive HSCORE, QIC SCORE, and PNA assays were extrapolated using DCC as the reference. ER immunodetection by PNA as compared with visual examination alone was enhanced by 18% (up to 88%) in sensitivity and 34% (up to 94%) in specificity, and the DCC concordance rate increased by 26% (up to 91%). A comparative chart extrapolating DCC from PNA was thus established.(ABSTRACT TRUNCATED AT 250 WORDS)     

Clinicopathological value of somatostatin type 2A and estrogen receptor immunoreactivity in human breast carcinoma

Somatostatin type 2A (sstr2A) and estrogen receptor (ER) are interrelated regulatory receptors present in normal breast epithelium and in a population of breast carcinomas. ER mediates growth stimulatory effects of estrogens whereas sstr2A mediates growth inhibitory actions of somatostatin. However, much work has been devoted to elucidate the biological role of ER, little is known about sstr2A in breast cancer. In the present study we examined immunoreactivity of sstr2A and ER in 64 breast carcinomas in correlation with tumor size and histological grade (HG), presence of lymph node metastasis (LNM), Nottingham prognostic index (NPI), and the patients' age. ER and sstr2A immunoreactivity were present in 78% and 63% of the breast carcinomas, respectively. Ninety percent of tumors immunoreactive for sstr2A were simultaneously immunoreactive for ER. ER immunoreactivity correlated significantly with lower HG (p=0.03) and better NPI (p=0.02). sstr2 A immunoreactivity correlated significantly with lower HG (p=0.012) but not with NPI (p=0.26). There was no correlation of sstr2A immunoreactivity and tumor size, patients' chronological age or LNM. The results confirm prognostic value of ER immunohistochemistry in breast carcinoma. However sstr2A cannot substitute ER for prognostic evaluation, sstr2A immunoreactivity being significantly associated with lower HG seems to represent an independent prognostic factor in breast carcinoma.