Immunohistochemical study of the ras oncogene expression in human breast lesions

An immunohistochemical study of ras oncogene expression in human breast lesions was carried out using a monoclonal antibody, Y13 259, to the ras encoded p21 protein. A total of 75 cases of breast disease examined included: 33 simple and complex cystic disease; 22 simple and hyperplastic fibroadenomas; 18 ductal, lobular and mixed carcinomas and 2 in situ carcinomas. Most of the complex cystic disease, hyperplastic fibroadenomas and all types of carcinomas showed high p21 expression as indicated by staining intensity. These results suggest that elevated ras expression may play an important role in the development of some premalignant and malignant breast lesions.     

Elevated expression of the myc gene in human benign and malignant breast lesions compared to normal tissue

Expression of the c-myc gene in human breast lesions and in adjacent normal tissue was studied by immunohistochemical analysis. The previously described monoclonal antibody Myc1-9E10 (1) which recognizes the p62 c-myc protein was used in paraffin tissue sections. A total of 101 cases of breast disease examined included 38 simple and complex cystic disease, 18 simple and hyperplastic fibroadenomas, 36 ductal and lobular carcinomas and 9 in situ carcinomas. Whereas the adjacent normal tissue was slightly positive, 25 out of 38 cystic disease, 7 out of 18 fibroadenoma, 36 out of 36 carcinoma and 9 out of 9 in situ carcinoma specimens showed moderate to high levels of p62 c-myc expression as indicated by staining intensity. These results suggest that the c-myc protein may play a role in breast neoplasia.     

HMGA1 protein expression in familial breast carcinoma patients

HMGA protein overexpression is associated with a highly malignant phenotype and it is also causally related to neoplastic cell transformation. Our previous results have shown that HMGA1 was not expressed in normal breast tissue whereas HMGA1 staining was intense in 25% of hyperplastic lesions with cellular atypia and in 60% of sporadic ductal carcinomas. Moreover, HMGA1 protein levels were significantly correlated with c-Erb-B2 expression. These results suggested HMGA1 expression as a novel prognostic factor in breast ductal carcinomas.In order to investigate whether the HMGA1 detection might have a prognostic role also for inherited breast carcinomas we have analysed the expression of the HMGA1 proteins in 116 breast familial carcinomas associated with BRCA1 or BRCA2 or negative for mutations in both genes (BRCAX). HMGA1 expression was weakly positive in 23 (20%), moderately positive in 34 (29%) and strongly positive in 20 (17%) breast carcinomas, and was not detected in 39 of them (34%). Statistical analysis of the immunostaining data showed that HMGA1 was significantly overexpressed, with a more intense staining, in BRCA2 (p = 0.0009) and BRCAX (p = 0.0134) patients compared to BRCA1 ones.Furthermore, in BRCA2 positive patients, the expression of HMGA1 seems to correlate with a favourable prognosis with a median overall survival of 65 months and a 5-year survival rate of 80% for HMGA1-negative patients, while median overall survival in the HMGA1-positive subsets was not reached with 5-year survival rates ranging from 84% to 100% of patients (p = 0.0198). Conversely, no correlation was found between HMGA1 expression and overall survival in patients carrying inherited mutations in the BRCA1 and in BRCAX patients.     

Immunohistochemical demonstration of metallothionein in normal human breast tissue and benign and malignant breast lesions

Metallothioneins (MTs) are a set of low molecular weight proteins with a high binding affinity to metal ions. MT over-expression has been recently demonstrated in invasive ductal carcinoma of the breast with poor clinical prognosis. In the present study, MTs have been immunohistochemically investigated in normal human breast tissue and a variety of benign, pre-invasive, and malignant breast lesions. In normal breast tissue, MTs were present in myoepithelial cells whereas the vast majority of luminal cells were MT negative. In lesions without increased cancer risk (adenosis and scleradenosis), MT was only immunolocalized in myoepithelial cells. In papillomas, MT was also found exclusively in myoepithelial cells. In most cases of epitheliosis, both the luminal and myoepithelial cells expressed MT. Atypicallobular hyperplasia, lobular carcinomain situ, and 13/15 invasive lobular carcinomas showed no MT over-expression. The two invasive lobular carcinomas with MT over-expression were classified as pleomorphic lobular carcinomas with apocrine differentiation. In contrast to lobular cancerization, 12/24 ductalin situ carcinomas and 9/20 invasive ductal carcinomas showed MT over-expression.In situ components found within invasive ductal carcinomas usually reflected the MT status of their invasive counterpart. It is concluded from our immunohistochemical results that breast carcinoma cases with MT overexpression arise from lesions which also show MT overexpression. Thus MT expression in carcinomas may be regarded as a genuine feature of the tumour cells and seems not to be related to endogenous or exogenous factors known to induce MT synthesis.     

bcl-2蛋白在良性乳腺病变及乳腺癌组织中的表达

目的　观察bcl-2蛋白在乳腺癌及良性乳腺病变中的表达。方法　采用免疫组化方法检测 15例正常乳腺组织、18例导管增生、10例不典型导管增生 ,10例小叶增生 ,6 9例浸润型乳腺癌和 45例乳腺导管原位癌中bcl -2蛋白表达。结果　bcl -2在 15例正常乳腺组织、18例导管增生、10例不典型导管增生及 10例小叶增生中均呈阳性 (10 0 % ) ,且均为高表达 ;bcl -2蛋白在 6 9例浸润型乳腺癌和 45例导管原位癌的表达阳性率分别为 6 5 2 % (45例 )和 75 6 % (3 4例 )。结论　bcl -2蛋白在正常乳腺组织良性乳腺病变的表达明显高于乳腺癌的表达。可能是在肿瘤的发展过程中bcl -2失去表达的结果 ,其原因和机理还有待进一步研究     

Clinicopathological significance of stromal myofibroblasts in invasive ductal carcinoma of the breast.

The purpose of this study was to investigate the distribution of CD34-positive fibroblasts and alpha-smooth muscle actin (alpha-SMA)-reactive myofibroblasts in the stroma of benign and malignant breast lesions and, secondly, to determine whether the presence of stromal myofibroblasts is associated with some of the clinicopathological characteristics of patients with invasive ductal carcinoma. The presence of stromal CD34-positive fibroblasts and myofibroblasts was investigated (as defined immunohistochemically) in 8 normal breast tissue samples, 58 invasive ductal carcinomas, 9 ductal carcinomas in situ and 16 specimens with benign lesions of the breast (fibroadenomas, ductal hyperplasias). We further studied the correlations between the presence of stromal myofibroblasts with 7 clinicopathological parameters in 58 invasive ductal carcinomas. The results indicated that the stroma of normal breast tissues contained CD34-positive fibroblasts. All benign breast lesions exhibited stromal CD34-positive fibroblasts. In contrast, the stroma of ductal carcinomas showed a complete loss of CD34-positive fibroblasts. alpha-SMA expression in stromal fibroblasts (myofibroblasts) was not detected in normal tissue samples or benign lesions except in 1 case of fibroadenoma, whereas positive myofibroblasts were found in 44.4% of ductal carcinomas in situ and 56.9% of invasive breast carcinomas. Comparison of clinicopathological parameters between invasive ductal carcinomas with and without stromal myofibroblasts revealed significant differences in lymph node metastasis, high histological grade and high microvessel density. These results suggest that CD34 loss and the presence of myofibroblasts favor the diagnosis of breast carcinoma. In invasive ductal carcinoma, the presence of stromal myofibroblasts correlated significantly with pathological parameters associated with a poor prognosis.     

Differential expression of cellular oncogenes in benign and malignant human breast tissue

We have examined 62 specimens of benign fibrocystic breast tissue, fibroadenomas, carcinomas and surrounding non-malignant tissue excised from 50 patients to determine the level of expression of 4 cellular oncogenes, c-myc, c-H-ras, c-K-ras, and c-N-ras. Our results demonstrate that in breast carcinoma the frequency and relative level of expression of these oncogenes are significantly greater than those found for benign breast tissue. However, some fibrocystic specimens having prominent hyperplastic features also exhibited enhanced oncogene expression. In view of the association between the increased frequency of carcinoma of the breast in women with a previous history of benign breast disease, it will be interesting to follow up donors of benign specimens to see if there is any relationship between the expression of oncogenes in such lesions and the development of carcinomas.     

Collagenase-3 expression in breast myofibroblasts as a molecular marker of transition of ductal carcinoma in situ lesions to invasive ductal carcinomas

Collagenase-3 (matrix metalloproteinase 13; MMP-13), a protease originally identified in breast carcinoma, is characterized by a potent degrading activity against a wide spectrum of extracellular matrix proteins. The aims of this study were to localize and identify the MMP-13-expressing cells in invasive human breast carcinoma and to evaluate the role of MMP-13 in transition to invasive lesions by studying ductal carcinoma in situ (DCIS). We found expression of MMP-13 in stromal fibroblast-like cells in all 21 invasive ductal carcinomas studied and in 4 of 9 invasive lobular carcinomas. In most carcinomas, expression of MMP-13 was limited to small stromal foci in the tumor area. Combined in situ hybridization and immunohistochemistry showed coexpression of alpha-smooth muscle actin immunoreactivity and MMP-13 mRNA in myofibroblasts. In contrast, cytokeratin-positive cancer cells, alpha-smooth muscle actin-positive vascular smooth muscle cells, CD68-positive macrophages, and CD31-positive endothelial cells were all MMP-13 mRNA negative. In situ hybridization for MMP-13 in 17 DCIS lesions revealed expression in 10 cases. Immunohistochemical analysis of all DCIS cases identified microinvasion in 8 of the 17 lesions. Seven of the eight lesions with microinvasion were MMP-13 positive. Further analysis showed that MMP-13 expression was often associated with the microinvasive events. This particular expression pattern was unique for MMP-13 among other MMPs analyzed, including MMP-2, -11, and -14. We conclude that MMP-13 is primarily expressed by myofibroblasts in human breast carcinoma and that expression in DCIS lesions often is associated with microinvasive events. On the basis of these data, we propose that MMP-13 may play an essential role during transition of DCIS lesions to invasive ductal carcinomas.     

Carcinomas arising in fibroadenomas: a report of two cases and a review of the literature

Carcinomas arising in fibroadenomas are rare. Two such cases are being presented; one is of the ductal variety and the other, lobular. Both cases clearly demonstrate the overall configuration of fibroadenomas with foci of in situ and infiltrating carcinoma, the surrounding breast tissue being devoid of malignant changes. The diagnosis was clinically unsuspected in both cases and in only one of them was it suspected at the time of gross pathological examination. The intent of this presentation is to increase general awareness as to the existence of carcinomas arising in fibroadenomas and also to actively discourage the practice of rendering gross pathological diagnoses of fibroadenomas at table diagnosis, however innocuous these neoplasms may appear to be, without microscopic examination by frozen sections.     

p63 expression in normal,hyperplastic and malignant breast tissues

BACKGROUND: p63 is a homologue of the p53 tumor suppressor gene and its protein is selectively expressed in the basal cells of a variety of epithelial tissues. It has recently been confirmed that p63 is expressed in the basal cells of normal prostate glands but not in prostatic carcinomas. Whether expression of p63 in breast correlates with tumor progression is the focus of this study. METHODS: Forty cases, which all contained normal breast tissue, ductal hyperplasia, ductal carcinoma in situ and invasive ductal carcinoma in the same patient were included in this investigation using an indirect immunohistochemical method and double staining. RESULTS: p63 was exclusively expressed in the myoepithelial cells of normal breast, partially expressed in ductal hyperplasia, rarely expressed in carcinoma in situ and not expressed in invasive carcinomas. CONCLUSION: The results suggest an association between loss of p63 expression and progression of breast ductal carcinoma. p63 immunostaining might be of assistance for distinguishing invasive ductal carcinoma from ductal carcinoma in situ or rare questionable ductal hyperplastic lesions, leading to correct therapy clinically.     

Nuclear morphometry of benign and malignant breast lesions

The mean nuclear area (MNA) of mammary gland epithelium was measured in 403 breast specimens, comprising 239 invasive carcinomas, 49 carcinomas in situ, 45 cases of fibrocystic disease (f.c.d.) with intraductal epithelial hyperplasia, and 60 cases of f.c.d. without intraductal hyperplasia. Normal breast tissue adjacent to other benign or malignant lesions was measured in 170 specimens. Statistical analysis revealed no difference between the MNA of invasive ductal carcinoma and ductal carcinoma in situ. The MNA of lobular and ductal carcinomas were significantly different. Significant differences were also found between ductal carcinoma and the two classes of f.c.d. The MNA of f.c.d. with and without intraductal hyperplasia were also significantly different, the former having the highest MNA. All breast lesions showed MNA significantly higher than that of normal breast epithelium. These findings show that there is a gradual increase in MNA from the baseline value of normal breast epithelium, via fibrocystic disease without and with intraductal proliferation to invasive carcinomas. Measurement of MNA may aid in pinpointing cases of intraductal epithelial hyperplasia with malignant potential.     

Differentiation between Cancerous and Normal Hyperplastic Lobules in Breast Lesions

Determining the risk that a particular area of hyperplastic breast tissue will progress to cancer is difficult and is currently expressed only as a general risk factor within the population. Using an antibody against the apoptotic purinergic receptor P2X7, we examined 40 cases each of the following histological categories: normal, moderate, florid and atypical hyperplasia, lobular carcinoma in situ, ductal carcinoma in situ, invasive lobular and invasive ductal carcinoma. These were previously diagnosed by H&E and supplied by clinical laboratories as tissue sections. Normal and mildly hyperplastic epithelium was devoid of the cytolytic P2X7 receptors whereas all epithelial cells in all cases of in situ or invasive lobular or ductal carcinoma labelled intensely. The lobular and ductal in situ cases labelled intracellularly while the invasive epithelial cancer cells showed intense cell surface label indicating an attempt was being made to induce apoptosis. All these receptors however are non-functional and thus unable to induce apoptosis. Approximately 10% of all hyperplastic lobules examined in the biopsied tissue, regardless of H&E classification, labelled for P2X7, which is suggestive of early metabolic cancerous change. The acini within lobules were either completely labelled with P2X7 or were completely devoid of the receptor. A potential advantage of this method lies in identifying early cancerous change in hyperplastic lobules and in establishing the true extent of cancerous spread in infiltrating lesions, thus facilitating the task of reporting clear surgical margins.     

Tissue binding of lectins in disorders of the breast

Twenty breast lesions including seven scirrhous ductal carcinomas, one infiltrating lobular carcinoma, one colloid carcinoma, four fibroadenomas, and seven cases of fibrocystic disease were analyzed by fluorescence microscopy for the presence and distribution of lectin-binding carbohydrates. Paraffin-embedded tissue sections were tested with wheat germ agglutinin (WGA), Ricin communis agglutinin I (RCA I), peanut agglutinin (PNA), Soybean agglutinin (SBA), Dolichos biflorus agglutinin (DBA), Ulex europaeus agglutinin I (UEA I), and concanavalin A (Con A). Brightest and most consistent staining regardless of the nature of the breast lesion was obtained with WGA followed in approximate order of staining intensity by RCA, PNA, SBA/DBA and Con A. UEA I stained many of the benign breast lesions but no malignant lesions. Lectin binding carbohydrate in benign lesions was localized mainly along the apices of mammary epithelial cells but there was considerable variation in staining patterns among malignant tumors. The fluorescence microscopic arrangement of lectin binding carbohydrate appears distinct for each malignant neoplasm of breast but is more consistent in benign conditions.     

Differential expression of UN1, early thymocyte-associated sialoglycoprotein,in breast normal tissue,benign disease and carcinomas

BACKGROUND: The UNI antigen (Ag) is a 120 kDa sialoglycoprotein which has been primarily found in human undifferentiated CD3dim thymocytes and leukemic T-cell lines, but subsequently also detected in solid tumors. We studied the expression of this Ag in a panel of normal and pathological breast tissues. MATERIALS AND METHODS: Analysis of UN1 Ag expression on tissue specimens was performed by immunohistochemistry and Western blotting. RESULTS: No Ag expression was found in 14 sections of normal tissue and 10 sections of benign nonproliferative lesions. Progressively increasing levels of UN1 Ag expression were found in fibroadenomas (24 positive out of 27 cases), proliferative lesions (9 cases), in situ (17 cases) and invasive carcinomas (56 cases). Finally, the highest expression was observed in 10 metastatic lesions. CONCLUSION: These data suggest that UN1 Ag is a promising marker of potential value for immunophenotyping studies and therapeutic applications in breast diseases.     

EGFR与HER-2在不同分子类型乳腺癌中的表达及其相关性分析

目的：探讨EGFR和HER-2在不同分子类型乳腺癌组织中的表达及其相关性。方法：回顾性分析2015年1月—2020年3月在邯郸市中心医院行手术治疗的乳腺癌患者资料650例(其中浸润性导管癌600例，浸润性小叶癌50例)及乳腺纤维腺瘤30例，应用免疫组织化学(IHC)方法对手术切除标本进行EGFR和HER-2蛋白表达检测，采用χ²检验及Spearman相关分析对数据进行统计学分析。结果：EGFR在乳腺浸润性导管癌中表达阳性率为48.7%，显著高于浸润性小叶癌(24.0%)及纤维腺瘤(16.7%)(P<0.01)。HER-2在乳腺浸润性导管癌中表达阳性率为27.8%，显著高于浸润性小叶癌(8.0%)及纤维腺瘤(6.7%)(P=0.002)。乳腺浸润性导管癌与浸润性小叶癌中，EGFR阳性率与HER-2评分等级呈正相关(r=1.000，P<0.05)，且EGFR阳性率与HER-2阳性表达呈正相关(r=1.000，P<0.05)。EGFR阳性率在HER-2过表达型与三阴型乳腺癌组织中显著高于Luminal A型和Luminal B型，且Luminal B中HER-2阳性显著高于HER-2阴性亚型(P<0.001)。结论：EGFR、HER-2阳性率在乳腺浸润性导管癌显著升高，且EGFR阳性率与HER-2的评分等级及阳性表达率均呈正相关，EGFR在HER-2过表达型与三阴型乳腺癌组织中显著升高，为进一步研究HER-2阳性乳腺癌及三阴型乳腺癌的靶向药物治疗提供线索及理论依据。     

Differential expression of keratins 13 and 16 in normal epithelium, benign lesions, and ductal carcinomas of the human breast determined by the monoclonal antibody Ks8.12

Previous electrophoretic analysis has indicated that keratins 13 and 16 (K13 and K16) are not present in normal human breast epithelium, but K16 is expressed in some ductal carcinomas (Moll et al., Cell, 31: 11-24, 1982). K16 may thus represent a marker for identifying a subset of ductal carcinomas and for distinguishing such tumor cells from normal cells. To explore this possibility further, we have used the monoclonal antibody Ks8.12, reportedly specific for K13 and K16 (Huszar et al., Differentiation, 31: 141-153, 1986), to examine keratin expression in human breast tissues. In immunocytochemistry the antibody reacted with epithelial cells of all normal samples, hyperplasias, and fibroadenomas. The staining was moderate to strong and always heterogeneous, involving most but not all luminal cells of ducts and acini. Myoepithelial cells were never stained. Of twenty-one ductal carcinomas examined, 90% gave a very weak or no reaction. The remaining 10% of cancers exhibited moderate to strong staining in about 50% of tumor cells. Cytoskeletal polypeptides extracted from the tissues and separated by polyacrylamide gel electrophoresis were analyzed by Western blotting to identify the polypeptides recognized by Ks8.12. In samples from normal tissue and a fibroadenoma, the antibody recognized a major Mr 48,000 component, a size appropriate for K16. In extracts from a hyperplasia and two of four carcinomas, the antibody detected a Mr 54,000 polypeptide, as well as a Mr 48,000 band, properties consistent with K13 and K16, respectively. Our results provide the first evidence indicating that K16 is present in normal as well as abnormal human breast epithelium. In addition, the data suggest that K13 may be expressed in some benign lesions and ductal carcinomas of the breast. Accordingly, Ks8.12 may prove to be useful for subclassifying ductal carcinomas and for discriminating between normal and certain benign and malignant disorders of human mammary epithelium.     

E-Cadherin Expression Correlates With Histologic Type But Not Tumour Grade in Invasive Breast Cancer

The traditional classification of infiltrating breast carcinomas into ductal and lobular can be diagnosticallychallenging in a small proportion of cases with equivocal histological features and in in-situ lesions withoverlapping features. Distinguishing between the infiltrating ductal (IDC) and lobular (ILC) carcinomas isclinically important because of the different pattern of systemic metastases and prognostic evaluation. E-cadherinis a potentially useful immunohistochemical marker which may serve to differentiate between the two tumourtypes. We therefore studied E-cadherin expression in 32 cases of breast carcinomas comprising 16 IDCs and 16ILCs. The correlation between E-cadherin expression and the histological grade of IDCs was also analysed. Ourresults showed complete loss of E-cadherin expression in all ILCs, while the IDCs consistently showed variableE-cadherin positivity. No significant correlation was found between E-cadherin expression and the histologicalgrade of IDCs. We conclude from this study that E-cadherin is a useful marker to differentiate between IDCand ILC of the breast. A larger study of IDCs is now needed to further evaluate the correlation between Ecadherinand tumour grade to estimate its prognostic potential.     

Telomerase activity in human breast cancer and benign breast lesions: Diagnostic applications in clinical specimens,including fine needle aspirates

We analysed telomerase activity in normal, benign and malignant breast tissues and in fine needle aspirates by a PCR-based assay. The tissue samples we used in this assay consisted of 20 cryostat sections, 10 μm thick, from each breast biopsy. This method was used to obtain effective extraction from small samples and to confirm the histological identity of the specimen by microscopical examination of serial sections. Fifty-two of 71 breast carcinomas were positive for telomerase activity, and the intensity of this was strong in most cases, whereas all 6 samples of normal breast tissue and 17 of fibrocystic disease were negative and only 1 of 15 fibroadenomas was positive. Invasive ductal carcinomas were more frequently positive than invasive lobular carcinomas. There was no correlation of telomerase activity with tumour size or the occurrence of lymph node metastasis. Evaluation of our assay system showed that a signal of telomerase activity was detectable in extracts from single cryostat sections (> 1 mm²) of a cancer specimen and from as few as 4 cells of a human breast cancer cell line. On the basis of the above data, we applied this assay to fine needle aspirates of breast lesions. Ten of 15 aspirates which had been cytopathologically diagnosed as cancer were strongly positive, while 26 of 29 benign aspirates were totally negative and the remaining 3 showed only borderline activity. In 3 cases, the telomerase result could have helped establish a diagnosis when the cytological observations were inconclusive. Our results indicate that this sensitive assay could become a useful new modality for supplementing microscopic cytopathology in the detection of cancer cells in small tissue biopsies and fine needle aspirates. © 1996 Wiley-Liss, Inc.