HER2 status in pure ductal carcinoma in situ and in the intraductal and invasive components of invasive ductal carcinoma determined by fluorescence in situ hybridization and immunohistochemistry

AIM: To determine the HER2 status of ductal carcinoma in situ (DCIS) and invasive ductal carcinoma (IDC) of the breast. The increased prevalence of HER2 amplification and overexpression in DCIS is considered to be maintained in the intraductal component of IDC; however, HER2 amplification and overexpression are detected much less in IDC. METHODS AND RESULTS: Fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) were performed to detect HER2 in 270 IDCs with an intraductal component and in 50 pure DCIS samples; IHC was also performed in 116 metastatic nodes. HER2 was found to be amplified in 77 cases (28.5%) and overexpressed in 79 (29.3%) of the 270 IDCs. HER2 amplification was similar between intraductal and invasive components of the same tumour. The concordance for HER2 status between invasive and intraductal components of individual tumours was 98.5% and 99.3% by FISH and IHC, respectively. HER2 was amplified in 25 (50%) of the 50 pure DCIS samples. HER2 overexpression in metastatic nodes resembled the HER2 status in the primary tumour for 108 (93.1%) of 116 cases (kappa =0.831). CONCLUSION: Our study indicates that the intraductal component of IDC may differ biologically when compared with pure DCIS. HER2 appears to lack a critical role in the progression from DCIS to IDC and HER2 status is maintained in metastatic lesions.     

乳腺小管癌与浸润性导管癌中NY-BR-1表达的对比观察

目的 探讨乳腺癌分化抗原NY-BR-1在乳腺小管癌中的表达,并与其在浸润性导管癌中表达进行对比.方法 收集29例小管癌和101例浸润性导管癌石蜡标本,采用免疫组化PV-9000通用型两步法分别进行NY-BR-1、ER、PR、Her-2、Ki-67、nm23、MDR-1及LRP的研究.结果 NY-BR-1在乳腺小管癌和浸润性导管癌中的表达率分别为51.7%、49.5%.NY-BR-1与组织学分化程度密切相关,其在浸润性导管癌组织学Ⅰ～Ⅲ级的阳性率逐渐降低分别为60%、52%、17%.NY-BR-1在小管癌中的表达明显高于浸润性乳腺癌组织学Ⅲ级的表达,差异有统计学意义(P<0.05).NY-BR-1在小管癌和Luminal A型乳腺癌中的表达无差异,在小管癌和浸润性导管癌组织学Ⅰ、Ⅱ级中的表达无差异.NY-BR-1与ER(rs=0.286,P=0.004)、PR(rs=0.252,P=0.010)的表达呈正相关.结论 作为分化抗原的一种,NY-BR-1在低分化的高级别癌中的表达率低,而在高分化的低级别癌中的表达率高.本研究结果显示NY-BR-1在小管癌和低级别浸润性导管癌中的表达率相似,提示小管癌与低级别、luminal型乳腺癌之间具有相似的组织学分化程度.     

乳腺浸润性导管癌HER2免疫组化检测结果判定的重复性研究

目的 探讨病理医师对乳腺浸润性导管癌HER2免疫组化(immunohistochemistry,IHC)检测结果判定的重复性及其影响因素,寻求提高判读重复性和准确性的措施.方法 参照<乳腺癌HER2检测指南>,2位乳腺专科病理医师共同读片选取56例浸润性导管癌的HER2 IHC切片作为研究对象.4位非专科病理医师独立盲法完成对该组病例IHC切片的前后两轮结果判定.对第一轮4位观察者间的判读重复性和观察者自身的判读重复性进行Kappa分析.同时将2位专科医师共同确定的判读结果作为HER2判读参考分级,4位非专科医师第一轮的判读结果分别与之进行配对Kappa分析.结果 观察者间的判读重复性为中等(K=0.593 9),重复性最高的级别是HER2为0(K=0.811 4),最低的是HER2为2 (K=0.473 6).判读级别简化为阳性和阴性后,观察者间的重复性明显提高(K=0.723 0).4位观察者自身的判读重复性极好(Kw=0.821 9～0.916 5),判读的准确性则与经验有关.结论 采用标准化的检测手段、深入学习和实践现有的判读标准以及联合判读等措施应有助于提高HER2 IHC判读的重复性和准确性.     

Expression of mucins (MUC1, MUC2, MUC5AC and MUC6) in mucinous carcinoma of the breast: comparison with invasive ductal carcinoma

AIMS: Mucinous carcinoma of the breast usually shows less frequent lymph node metastasis and more favourable outcome compared with invasive ductal carcinoma. The aim of this study is to compare the expression profiles of several mucins in mucinous carcinomas and invasive ductal carcinomas to gain insight into the relationship between the less aggressive biological nature of mucinous carcinoma and the role of mucins. METHODS AND RESULTS: We examined the expression profiles of MUC1 (membrane-bound mucin) of different glycoforms (from non-glycosylated form to fully glycosylated form), MUC2 (intestinal type secretory mucin), MUC5AC (gastric surface type secretory mucin) and MUC6 (gastric pyloric gland type secretory mucin) in 17 mucinous carcinomas and 46 invasive ductal carcinomas using immunohistochemistry. Various glycoforms of MUC1 were expressed frequently in both mucinous carcinomas (65-100%) and invasive ductal carcinomas (92-100%), although non-glycosylated MUC1 (MUC1/CORE) and fully glycosylated MUC1 (MUC1/HMFG-1) showed significantly lower expression rates in mucinous carcinomas compared with those in invasive ductal carcinomas. The expression rates of MUC2 (94%) and MUC6 (71%) in mucinous carcinomas were significantly higher than those of MUC2 (15%) and MUC6 (15%) in invasive ductal carcinomas. There was no significant difference in the expression rate of MUC5AC in mucinous carcinomas (12%) and that in invasive ductal carcinomas (4%). CONCLUSIONS: The expression rate of MUC1/CORE and MUC1/HMFG-1, which is related to poor prognosis in the gastric and colorectal cancers, is low in mucinous carcinomas. The high expression rate of gel-forming secretory mucins (MUC2 and MUC6) in mucinous carcinoma suggests that high production of these types of mucins may act as a barrier to cancerous extension resulting in their less aggressive biological behaviour.     

Membranous and cytoplasmic staining of Ki67 is associated with HER2 and ER status in invasive breast carcinoma

Aims:  Membranous and cytoplasmic Ki67 immunoreactivity has recently been observed in a number of histopathological entities, but frequency of occurrence and relationship to prognosis in more common cancers have not been described. The aim was to describe the pattern and frequency of membranous/cytoplasmic Ki67 in a cohort of invasive breast carcinomas, and their associations with grade, HER2 amplification and oestrogen receptor (ER) expression.
Methods and results:  Three hundred and twenty-two cases of invasive ductal carcinoma were assessed for histological grade, Ki67 (MIB-1 clone) proliferation index and pattern of immunoreactivity, ER expression by immunohistochemistry, and HER2 amplification status by fluorescence in situ hybridization. Overall, 26/322 (8%) breast carcinomas showed membranous/cytoplasmic Ki67, and expression was significantly associated with grade 3, HER2-amplified and ER− tumours. Membranous/cytoplasmic Ki67 was not, however, an independent prognostic factor on multivariate analysis.
Conclusions:  Membranous/cytoplasmic Ki67 identifies a group of breast carcinomas that may be important to consider separately in prognostic and predictive studies. The mechanism of subcellular Ki67 relocalization remains elusive and further studies are required to establish both the cause and effect of this unusual pattern of Ki67 immunoreactivity.     

乳腺浸润性导管癌微卫星不稳定性

目的 探讨乳腺浸润性导管癌的微卫星不稳定(MSI)性及其与临床病理资料的关系。方法 选取10个微卫星位点，从石蜡包埋的存档标本中选取34例肿瘤组织和其对应的自身正常对照组织，提取DNA后用PCR扩增，6％聚丙稀酰胺凝胶电泳，银染显色后进行微卫星不稳定性分析。用免疫组化S-P法观察p53、c-erbB-2、PR、ER在乳腺癌中表达情况。结果 在34例乳腺浸润性导管癌中有9例(26．47％)至少1个位点出现MSI。MSI和病人年龄、肿瘤大小、病理分级、淋巴结转移、p53、c-erbB-2之间没有明显的相关性。但ER和PR阴性的病例出现MSI的比例远远高于ER和PR阳性病例。结论在乳腺癌的发生、发展过程中出现MSI,并可能和ER、PR的表达降低有关。     

乳腺癌组织中Caveolin-1、MMP-2的表达及意义

目的 探讨Caveolin-1及MMP-2蛋白在乳腺癌及正常乳腺组织中的表达及两者的相关性.方法 采用EnVision法免疫组化检测86例乳腺癌及20例正常乳腺组织中Caveolin-1及MMP-2蛋白的表达.结果 乳腺癌中Caveolin-1的阳性率(52.3%)低于正常乳腺组织(95.0%)(P<0.05),而MMP-2的阳性率高于正常乳腺组织(P<0.05).Caveolin-1在乳腺癌的表达与淋巴结转移、临床分期相关(P<0.05),而与患者年龄、组织学分级及肿瘤大小无关(P>0.05).MMP-2在乳腺癌的表达与淋巴结转移、组织学分级及临床分期相关(P<0.05),而与患者年龄及肿瘤大小无关(P>0.05).在乳腺癌中Caveolin-1和MMP-2的表达呈负相关(r=-0.472,P<0.05).结论 Caveolin-1低表达与MMP-2过表达可能是乳腺组织恶性转变以及乳腺癌发生浸润转移的重要生物学标志,联合检测Caveolin-1和MMP-2对预测乳腺癌的浸润转移有重要意义.     

Cytological criteria for the diagnosis of intraductal hyperplasia, ductal carcinoma in situ, and invasive carcinoma of the breast

The advent of mammography screening presents a diagnostic challenge to the cytopathologist as an increasing proportion of breast lesions requiring investigation will be nonpalpable and up to 40% will be accounted for by atypical intraductal hyperplasia and ductal carcinoma in situ, as opposed to previously, when these lesions represented less than 10% of palpable tumors. We studied 133 fine-needle aspirates from breast tumors and found that nuclear morphology, myoepithelial cells, signs of invasion, and degree of cellular dissociation are among the most potent factors discriminating between benign epithelial proliferations, atypical intraductal hyperplasia, ductal carcinoma in situ, and invasive carcinoma.     

Low expression of HER2 protein in breast cancer is biologically significant

HER2 status is routinely tested using immunohistochemistry or FISH following the licensing of a therapeutic agent targeting HER2. However, neither of these methods provides quantitative information relating to the 70-80% of patients with levels of expression lower than the assay detection thresholds. In this study, radioimmunohistochemistry was used to detect quantitative HER2 protein expression in 178 breast cancers. Survival analysis was performed, as were correlations with known prognostic variables and with overexpression of other HER family members. It is demonstrated that the populations expressing very high and very low levels of HER2 are each associated with increased risk of cancer-specific death on survival analysis (p = 0.0043). The group with low levels of HER2 was more likely to be of higher grade, EGFR-positive and ER/HER3/HER4-negative. HER2-positive cases were frequently ER-negative/HER3-positive, whilst cases with normal HER2 expression were often ER-positive/HER4-positive. The aggressive nature of the tumour group with low HER2 expression may be explained by actions of other HER family members, particularly EGFR, but whether these or other factors have a negative regulatory effect on HER2 expression remains to be determined.     

The expression pattern of MUC1 (EMA) is related to tumour characteristics and clinical outcome of invasive ductal breast carcinoma

AIMS: To clarify MUC1 patterns in invasive ductal breast carcinoma and to relate them to clinicopathological parameters, coexpression of other biological markers and prognosis. METHODS AND RESULTS: Samples from 243 consecutive patients with primary ductal carcinoma were incorporated into tissue microarrays (TMAs). Slides were stained for MUC1, oestrogen receptor (ER), progesterone receptor (PR), Her2/neu, p53 and cyclin D1. Apical membrane MUC1 expression was associated with smaller tumours (P = 0.001), lower tumour grades (P < 0.001), PR positivity (P = 0.003) and increased overall survival (OS; P = 0.030). Diffuse cytoplasmic MUC1 expression was associated with cyclin D1 positivity (P = 0.009) and increased relapse-free survival (RFS; P = 0.034). Negativity for MUC1 was associated with ER negativity (P = 0.004), PR negativity (P = 0.001) and cyclin D1 negativity (P = 0.009). In stepwise multivariate analysis MUC1 negativity was an independent predictor of both RFS [hazard ratio (HR) 3.5, 95% confidence interval (CI) 1.5, 8.5; P = 0.005] and OS (HR 14.7, 95% CI 4.9, 44.1; P < 0.001). CONCLUSIONS: The expression pattern of MUC1 in invasive ductal breast carcinoma is related to tumour characteristics and clinical outcome. In addition, negative MUC1 expression is an independent risk factor for poor RFS and OS, besides 'classical' prognostic indicators.     

Expression and mutational status of RON in neoplastic lesions of the breast: analysis of MSP/RON signaling in ductal carcinoma in situ and invasive ductal carcinoma

Recepteur d'origine nantais (RON) is a receptor tyrosine kinase closely related to MET and involved in tumorigenesis. We investigated the roles of aberrations in RON and its ligand, macrophage-stimulating protein (MSP), in invasive ductal carcinoma (IDC, n = 81), ductal carcinoma in situ (DCIS, n = 26), and in benign lesions (n = 20) of mammary gland. Expression of RON and MSP was evaluated by immunohistochemistry and the mutational status of a region containing the proteolytic cleavage site in exon 1 and each exon of the kinase domain (exon 14-20) of RON was screened by polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) analysis. The proportion of cases positive for RON expression was significantly different between malignant [86% (92/107)] and benign [40% (8/20)] lesions. RON expression was positive in both IDC and DCIS [90% (73/81) and 73% (19/26), respectively], whereas MSP expression was present in 54% (44/81) of IDC and absent in DCIS. RON expression correlated significantly with the histological grade of DCIS. No mutations were detected in the examined regions of RON in breast cancer samples as confirmed by PCR-SSCP. The findings suggest the involvement of RON expression in the development of breast cancer, and that an autocrine/paracrine loop of RON seems to affect tumor invasiveness.     

Notch1 single nucleotide polymorphism rs3124591 is associated with the risk of development of invasive ductal breast carcinoma in a Chinese population

Accumulated evidence has revealed the presence of Notch receptor polymorphisms in non-tumorous diseases; however, few studies have investigated the association of Notch polymorphisms with breast cancer risk. A total of 100 invasive ductal carcinoma (IDC) and 50 ductal carcinoma in situ (DCIS) patients and 100 usual ductal hyperplasia (UDH) controls were genotyped for the following Notch receptor single nucleotide polymorphisms (SNPs) using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry: Notch1, rs3124591; Notch2, rs11249433; Notch3, rs3815188, and rs1043994; and Notch4, rs367398, and rs520692. Immunohistochemistry was used to determine the effect of Notch polymorphisms on corresponding Notch protein expression in successfully genotyped patients. The frequency of rs3124591 TC genotype was significantly higher in IDC (24.7%, 20/81) and DCIS (30%, 12/40) patients than in UDH controls (8%, 8/97) (P = 0.002 and P = 0.011, respectively). However, the distribution of other SNP genotypes was not significantly different between IDC and DCIS patients and UDH controls. The frequency of TC genotype was significantly higher in poorly differentiated tumors than in well-differentiated and moderately differentiated tumors (P = 0.022). Importantly, a positive correlation between the rs3124591 TC genotype and high Notch1 protein expression was observed in DCIS patients (P = 0.043) but not in IDC patients. This is the first study to suggest an increased risk of IDC and DCIS of the breast for the Notch1 rs3124591 variant. Furthermore, given the inconsistent associations between the rs3124591 variant and Notch1 expression in IDC and DCIS, this variant may affect breast cancer risk through mechanisms in the latter stage other than alterations in Notch1 protein expression.     

Concordance of HER2 status assessed on needle core biopsy and surgical specimens of invasive carcinoma of the breast

Lee A H S, Key H P, Bell J A, Hodi Z & Ellis I O
(2012) Histopathology  60, 880–884 Concordance of HER2 status assessed on needle core biopsy and surgical specimens of invasive carcinoma of the breast Aims: Human epidermal growth factor receptor 2 (HER2) status of invasive breast cancers is vital for selection of patients for trastuzumab treatment. This study aimed to assess the level of agreement of HER2 status in core biopsy and excision specimens using immunohistochemistry, with in‐situ hybridisation for equivocal cases. Methods and results: 300 consecutive invasive carcinomas with core biopsy and surgical specimens had HER2 assessed on both specimens. Immunohistochemistry was performed first. Fluorescence in‐situ hybridization (FISH) was automatically performed if the immunohistochemistry was scored as equivocal (2+). There was agreement between the HER2 status of the two specimens in 294 tumours (98%). In two carcinomas the core was negative and the excision specimen showed focal strong staining with amplification. In four carcinomas the core biopsy was negative and the excision showed 2+ staining with amplification in at least one block (although in three there was no amplification in a second block). Conclusion: There is excellent agreement between HER2 assessed in core biopsy and surgical specimens. Discrepancies arise most frequently due to focal amplification or levels of gene amplification around the cut‐off for defining positivity.     

Association of ultrasonographic features with NGX6 expression and prognosis in invasive ductal breast carcinoma

Introduction: Nasopharyngeal carcinoma-associated gene 6 (NGX6) is a newly discovered tumor suppressor gene. It contains one epidermal growth factor (EGF)-like domain. Many studies have shown that proteins contain (EGF)-like domain structure affect a variety of biological actions. However, little is known about the relationships between NGX6 expression and biological behaviors in invasive ductal breast carcinoma (IDBC). The study discussed the association of ultrasonographic features with NGX6 expression and prognosis in IDBC. Methods: Ultrasonographic feature and clinical data in 122 patients with IDBC were retrospectively analyzed. NGX6 expression of IDBC was measured using immunohistochemistry methods. Results: The incidence of the burr sign, lymph node metastasis and abundance blood flow in NGX6 expression negative groups were higher than those in positive groups. Kaplan-Meier analysis showed that the association between NGX6 positive expression and higher disease-free survival (DFS) or higher overall survival (OS); Lymph node metastasis is associated with lower DFS or lower OS; Lower blood flow grade is associated with higher DFS. In univariate and multivariate survival analysis, NGX6 expression, lymph node metastasis, TNM and the blood flow grade were the independent prognostic factors for DFS and OS of IDBC. Conclusions: ultrasonographic features are associated with NGX6 expression in IDBC. NGX6 is involved in the invasion and metastasis activity of IDBC. Our results suggest that NGX6 may be employed as a promising prognostic factor and useful therapeutic target for IDBC. Combination of ultrasonic findings and NGX6 detection may yield clinically useful information for IDBC prognosis.     

乳腺癌Her2基因荧光原位杂交及其临床病理关系

目的探讨Her2基因过表达的比例及其与临床病理之间的联系。方法收集乳腺浸润型导管癌标本50例,总结其临床及病理资料,应用石蜡包埋组织切片,以EnVision两步法进行免疫组织化学染色标记ER、PR及HER2,以金菩嘉DNA探针试剂盒行荧光原位杂交检测HER2基因。结果患者平均年龄55.5岁,病理组织分级Ⅰ级11例,Ⅱ级30例,Ⅲ级9例。术后分期Ⅰ期13例,ⅡA期15例,ⅡB期13例,ⅢA期6例,ⅢB期2例,ⅢC期1例。淋巴结中位转移率6.91%。ER阳性33例,PR阳性32例,IHC法HER2阳性40例,FISH阳性33例,阴性17例。FISH法检测HER2基因与HER2蛋白过表达之间相关(P0.05),其与病理分级、术后分期、淋巴结转移率及ER、PR的表达无相关(P0.05),3例脉管内瘤栓及3例原发瘤数大于1者,FISH均为阳性。结论FISH检测HER2基因扩增和免疫组化检测HER2蛋白一致性较好,HER2基因与脉管内瘤栓及多个原发瘤有一定关系,但其作为间接预测预后的指标有待于进一步研究。     

Expression patterns of stromal MMP‐2 and tumoural MMP‐2 and ‐9 are significant prognostic factors in invasive ductal carcinoma of the breast

Matrix metalloproteinases (MMPs) are matrix‐degrading enzymes that play a pivotal role in aggressive behaviours, such as rapid tumour growth, invasion, and metastasis, of several types of solid tumours. In particular, stromal MMP‐2 plays important roles in the progression of malignant tumours, but most clinical studies have focused on tumoural MMP‐2 and ‐9 expression, and not stromal MMP‐2 expression. One hundred and seventy‐seven cases diagnosed as invasive ductal carcinoma of the breast between 2000 and 2005 were included in this study. Expressions of tumoural MMP‐2 and ‐9 and stromal MMP‐2 were analysed by immunostaining on a tissue microarray. Subsequently, the associations between those results and various clinicopathological parameters were evaluated. Stromal MMP‐2 expression correlated significantly with clinicopathological parameters such as advanced T category, larger tumour size, high histological grade, tumour necrosis, ER‐ and PR‐negative, and HER‐2‐positive (all p < 0.05). In univariate and multivariate analyses, overall survival was linked with stromal MMP‐2 expression as well as dual expression of stromal MMP‐2 and tumoural MMP‐2 and ‐9 (all p < 0.05). Stromal MMP‐2 expression may play a crucial role in predicting aggressive clinical behaviour in breast cancer patients.     

Expression of MT-1 MMP, MMP2, MMP9 and TIMP2 mRNAs in ductal carcinoma in situ and invasive ductal carcinoma of the breast

We investigated the expression of membrane type-1 (MT1)-MMP, MMP2, MMP9 and TIMP2 mRNAs and their roles in ductal carcinoma in situ (DCIS) and T1 and T2 invasive ductal carcinoma of the breast. We further compared these two types of carcinomas for differences in microvessel density, and expression of angiogenic factors and CD44std. MT1-MMP, MMP2, MMP9 and TIMP2 mRNA were expressed in both DCIS and invasive ductal carcinomas. Expression rates of MT1-MMP, MMP2, MMP9 and TIMP2 mRNAs were not statistically different between DCIS and invasive ductal carcinomas, nor did they differ statistically when grouped by tumor size, histologic grade or nuclear grade of invasive ductal carcinoma. Microvessel density and expression of VEGF and TGF-beta1 were not statistically different between DCIS and invasive ductal carcinoma. CD44std expression was significantly increased in DCIS compared to invasive ductal carcinoma (p < 0.05) and it was also significantly increased in lower clinical stage, histologic grade and nuclear grade of invasive ductal carcinoma (p < 0.05). Axillary node metastasis was significantly correlated with MT1-MMP mRNA, VEGF and TGF-beta1 expression (p < 0.05) and MT1-MMP mRNA was positively correlated with VEGF expression and TIMP2 mRNA (p < 0.05). In summary, patterns of MMP mRNA expression in DCIS and invasive ductal carcinoma suggest that the invasive potential of breast carcinoma is already achieved before morphologically overt invasive growth is observed. As MT1-MMP mRNA expression is significantly correlated with axillary nodal metastasis, it may be useful as a prognostic indicator of invasive ductal carcinoma. Considering the positive correlation of MT1-MMP mRNA and TIMP2mRNA expression, our finding supports a role for TIMP2 in tumor growth, as well as the utility of CD44std as a prognostic indicator of breast cancer.