乳腺癌ER,PR状态与细胞超微结构变化的形态定量分析

用免疫组化ＰＡＰ法检测３０例乳腺癌雌激素受体（ＥＲ）和孕激素受体（ＰＲ），从中选取ＥＲ、ＰＲ阳性者（Ｅ＾＋Ｐ＾＋）６例，阴性者（Ｅ＾－Ｐ＾－）５例作透射电镜观察，对部分细胞器连接的变化进行形态定量分析。结果Ｅ＾－Ｐ＾－组癌细胞内线粒体、粗面内质网及溶酶体的含量明显高于Ｅ＾＋Ｐ＾＋组，且差异有显著性（Ｐ＜０．０５，０．０１，０．０１）；Ｅ＾－Ｐ＾－组细胞间的桥粒及镶嵌连接有减少趋势，但差异无显著性     

Immunohistochemical evaluation of human epidermal growth factor receptor 2 and estrogen and progesterone receptors in invasive breast cancer in women

Introduction

Estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER-2) expression are crucial in the biology of breast carcinoma. HER-2/neu gene is amplified and overexpressed in 15-30% of invasive breast cancers. HER-2-positive breast cancers have worse prognosis than HER-2 negative tumors and possess distinctive clinical features. The aim of this study was to assess the expression of HER2 in cancer tissue of patients with invasive breast cancer in correlation with tumor type, histological grade, tumor size, lymph node status, and expression of estrogen receptor and progesterone receptor.

Material and methods

Formalin-fixed, paraffin-embedded tissues from 40 patients with invasive HER-2-positive breast cancer and from 191 patients with HER-2-negative breast cancer were used in this study. HER2 expression was determined using the test HerceptTest™ DAKO.

Results

Among 231 cases of breast cancer, 18 invasive lobular carcinomas and 213 invasive ductal carcinomas were diagnosed. Sixty percent of HER-2-positive breast cancers were ER-positive compared with 77% in the HER-2-negative group (p = 0.002). The expression of PR was observed in 43% of HER-2-positive breast cancers and in 72% of HER2-negative tumors (p = 0.003). Excessive expression of HER2 protein was detected in 60% of patients positive for estrogen receptors, which may worsen prognosis in these patients.

Conclusions

Determination of HER2 overexpression in breast cancer patients, allows for a determination of a group of patients with a worse prognosis.     

Expression of estrogen and progesterone receptors in cytologic specimens using various fixatives

Estrogen and progesterone receptor reactivity may be useful in identifying possible primary sites of metastatic disease or directing therapy in tumors of the female genital tract, including breast, ovary, and endometrium. Various methods have been described for the immunocytochemical evaluation of estrogen receptor (ER) and progesterone receptor (PR) status of cytologic specimens but our results have been variable. We evaluated the effectiveness of various fixatives [cytospin collection fluid, Shandon, Pittsburgh, PA (SH); ethanol (ETH); and formalin (FOR)] for fixation of smears (SM) and cell block (CB) material. The percentage and intensity of tumor nuclei of SM, CB, and tissue sections (TS) stained for ER and PR by the avidin-biotin-peroxidase complex technique were compared. Samples were considered ER or PR positive when ≥20% of tumor nuclei were stained. The sensitivity of ER analysis of SMs and CBs in each fixative compared to formalin-fixed paraffin-embedded tissue sections were as follows, SM (SH) 88%, SM (ETH) 14%, CB (SH) 58%, CB (ETH) 43%, and CB (FOR) 70%. The sensitivity of PR determination on SMs and CBs was SM (SH) 71%, SM (ETH) 6.0%, CB (SH) 25%, CB (ETH) 33%, CB (FOR) 80%. These findings indicate that of the fixatives evaluated for ER analysis SMs fixed in SH provided the best results. For PR evaluation, CBs fixed in FOR gave the best results. Diagn Cytopathol 1996;15:78–83. © 1996 Wiley-Liss, Inc.     

Endometrial progesterone and estrogen receptors and bleeding disturbances in depot medroxyprogesterone acetate users

BACKGROUND: Depot medroxyprogesterone acetate (DMPA) is a very popular hormonal contraceptive. Unpredictable bleeding disturbances are the main reasons for discontinuation and may be mediated through endometrial hormone receptors. This study was aimed to compare the expression of progesterone and estrogen receptors in the endometrium of bleeding DMPA users with that of amenorrhoeic DMPA users. METHODS: Subjects were recruited between April 2000 and January 2001. On the day of the third DMPA injection, 42 amenorrhoeic DMPA users and 42 DMPA users who had frequent or prolonged endometrial bleeding and were bleeding on that day were matched by age and body mass index. Endometrial biopsies were collected for immunohistochemical study of progesterone receptor A plus B (PRAB) and B alone (PRB) and estrogen receptor alpha (ERalpha) and beta (ERbeta) expression. RESULTS: There were 23 adequate endometrial samples from each group. There were no differences in any of a series of comparisons of PRAB, PRB, ERalpha and ERbeta expression in glands or stroma between the groups. Serum estradiol and progesterone levels were also not different between the groups. CONCLUSIONS: Endometrial PRAB, PRB, ERalpha and ERbeta expression in glands and stroma was not different between DMPA users who had frequent or prolonged bleeding and amenorrhoeic DMPA users.     

子宫腔粘连患者子宫内膜组织中ER、PR的表达及意义

目的：探讨雌激素受体( ER)、孕激素受体( PR)在子宫腔粘连( intrauterine adheious, IUA)患者子宫内膜组织中的表达及其临床意义。方法采用免疫组化MaxVision两步法和实时荧光定量PCR( qRT-PCR)技术检测IUA组(研究组)和非IUA组(对照组)子宫内膜组织中ER、PR的表达水平。结果免疫组化检测显示ER蛋白在研究组(3．52±0．71)和对照组(2．75±1．00)中的表达差异有统计学意义(P=0．01);qRT-PCR法检测显示ER mRNA在研究组(1．59±0．26)和对照组(1．00±0．19)中的表达差异有统计学意义(P=0．00)。免疫组化检测显示PR蛋白在研究组(3．26±0．70)和对照组(3．58±0．28)中的表达差异无统计学意义(P=0．12);qRT-PCR法检测显示PR mRNA在研究组(1．15±0．21)和对照组(1．00±0．31)中的表达差异无统计学意义(P=0．21)。免疫组化检测显示ER、PR蛋白分别在子宫内膜腺体和子宫内膜间质中均有表达,差异有统计学意义(χ2=5．797,P=0．016;χ2=4．857,P=0．027)。结论子宫内膜组织中ER表达研究组高于对照组,PR在两组之间的表达无差异;ER、PR蛋白在两组子宫内膜腺体中表达较多,在子宫内膜间质中表达较少甚至不表达;ER、PR表达的特点为IUA患者临床使用雌、孕激素治疗提供一定的理论参考。     

抗PR,抗ER单克隆抗体的制备及鉴定

目的自行研制抗人孕激素受体（ＰＲ）和抗人雌激素受体（ＥＲ）单克隆抗体（ＭｃＡｂ）,以期达到可替代国外ＭｃＡｂ之目的。方法以ＰＲ氨基端基因表达蛋白产物及ＥＲＤ合成肽为抗原,制备小鼠抗ＰＲ、抗ＥＲＭｃＡｂ,采用Ｗｅｓｔｅｒｎｂｌｏｔ,免疫组化ＳＰ法,免疫荧光技术和流式细胞仪鉴定自制ＭｃＡｂ的性质。结果（１）抗ＰＲＭｃＡｂ与ＰＲ阳性乳腺癌细胞Ｔ４７Ｄ的提取物,抗ＥＲＭｃＡｂ与ＥＲ阳性之ＭＣＦ７细胞的提取物转移于硝酸纤维膜上的蛋白显示特异反应带;（２）抗ＰＲ、抗ＥＲＭｃＡｂ分别检测４０和３５例乳腺癌组织石蜡切片,染色效果与Ｚｙｍｅｄ公司生产的ＰＲ、ＥＲＭｃＡｂ相当,阴、阳性符合率均为１００％。免疫组化与ＤＣＣ法一一对应比较,符合率分别为８５０％、９１４％。（３）荧光显微镜和激光共聚焦扫描显微镜观察荧光标记阳性反应物主要位于胞核。（４）流式细胞仪检测,荧光指数（Ｆ１）＞１．０,阴性对照Ｆ１＜１．０。结论自制抗ＰＲ、抗ＥＲＭｃＡｂ特异性强,灵敏度高,结果与Ｚｙｍｅｄ公司产品相同,操作简便,适于临床应用     

New monoclonal antibodies to oestrogen and progesterone receptors effective for paraffin section immunohistochemistry

Assessment of oestrogen and progesterone receptors (ER and PgR) in breast cancer is widely used for the prediction of response to endocrine therapy and as a prognostic marker. Cytosolic assays have been replaced in many centres by immunochemical techniques, which have many advantages including applicability to small samples, simplicity, and cost-effectiveness. This study describes the generation and characterisation of two novel murine monoclonal antibodies recognizing ER and PgR, designated NCL-ER-6F11 and NCL-PGR respectively, which are effective in heat-treated formalin-fixed, paraffin-embedded tissue. The antibodies have been characterized by Western blotting and by immunohistochemistry on normal and pathological breast and other tissues. NCL-ER-6F11 has been shown to compare favourably with a currently available ER antibody. These antibodies may prove of value in the assessment of hormone receptor status in human breast cancer. © 1997 John Wiley & Sons, Ltd.     

Estrogen and progesterone hormone receptor status in breast carcinoma: comparison of immunocytochemistry and immunohistochemistry

We evaluated the correlation between histologic and cytologic specimens in the determination of estrogen receptor (ER) and progesterone receptor (PR) status in breast carcinoma and investigated the causes of clinically significant discrepancies. We analyzed 70 immunoassays for ER and 60 for PR from 71 patients with breast carcinoma. Concordance between cytology and histology was 89% for ER and 63% for PR using scores from pathology reports. Concordance between cytology and histology was 98% for ER and 91% for PR using consensus scores (obtained after reevaluation by the team of pathologists). Thirty of 130 (23%) tests had clinically relevant discrepancies, 53% of which were caused by wrong interpretation of cytologic findings, 10% by wrong interpretation of histologic findings, 17% by sampling error and 20% were not available for reevaluation. Wrong interpretation of the results for ER and PR status in cytology was a far more frequent cause of clinically relevant discrepancies than sampling errors. The use of strict criteria is recommended.     

Demonstration of oestrogen and progesterone receptors in freeze-dried, paraffin-embedded sections of breast cancer

Immunohistochemical evaluation of oestrogen and progesterone receptors is of importance in evaluating human breast tumours. Staining techniques can be performed on snap-frozen, cryostat-cut tissues or, as recently reported, on formalin-fixed, paraffin-embedded tissues. These methods are, however, limited by several drawbacks, including difficulties in retrospective studies and in storage of the material, and the relatively high frequency of false negative results for chemically fixed specimens. We therefore investigated the application of freeze-drying technology to assess the feasibility and reliability of this technique as an alternative method for diagnostic breast pathology. Morphological and immunohistochemical studies were performed on snap-frozen, freeze-dried and paraffin-embedded tissue obtained from 16 cases of benign and malignant breast neoplasms. Our results showed good preservation of tissue morphology, similar to standard formalin fixation, and excellent preservation of antigenic reactivity of nuclear receptors, comparable to that obtained with cryostat sections. We therefore suggest that freeze drying and paraffin embedding of frozen tissue blocks is equivalent or even preferable to formalin fixation for the demonstration of oestrogen and progesterone receptors, at least in the case of small tumours.     

Estrogen receptor β– an independent prognostic marker in estrogen receptor α and progesterone receptor‐positive breast cancer?

Both subtypes of estrogen receptor (ER), ERα and ERβ, are normally present in the mammary gland. The role of ERα as a prognostic marker in breast cancer is well established due to the beneficial effect of providing tamoxifen as adjuvant therapy. The role of ERβ, however, is less clear. To gain insight into the importance of ERβ in breast cancer, 145 primary breast cancers were examined by immunohistochemistry for ERβ, and the expression level was compared with ERα and progesterone receptor (PR) status. Especially, we wanted to examine the significance of ERβ in the contrasting ERα+/PR+ and ERα?/PR? subgroups. In the ERα+/PR+ subgroup (dual positive), the survival difference between patients with low, medium and high ER β level was statistically significant (p = 0.004), with more than 70% of patients with medium and high ERβ levels surviving 100 months, compared with less than 30% in the group with low ERβ level. Further, for ERα+/PR+ patients there was a reduced risk of fatal outcome by multivariate analysis with increasing ERβ levels (p(trend) < 0.01 [univariate analysis]; p(trend) = 0.05 [multivariate analysis]). The risk was 31% and 27% for medium and high ERβ levels, respectively, compared with low ERβ level, adjusting for standard prognostic factors such as tumor diameter, nuclear tumor grade (quantified by mean nuclear area), lymph node status, and patient age at operation. For patients with ERα?/PR? tumors (dual negative), however, there was no association between ERβ levels and patient outcome. Our findings indicate that ERβ expression provides independent prognostic information for breast cancers with ERα/PR‐positive status, a feature typical among screen‐detected breast cancers. The role of ERβ needs to be further evaluated especially in this group of breast cancers.     

p53 expression in human breast cancer related to survival and prognostic factors: an immunohistochemical study

In a study of 90 breast cancer patients, tumour p53 protein expression was determined by immunohistochemistry using the monoclonal antibody PAb1801. Patient lymph node status and Bloom's grade were determined, and both oestrogen and progesterone status assessed, also by immunohistochemistry. Lymph node status, tumour grade, and progesterone receptor status all had a significant influence on survival. Patients with p53-positive tumours showed poorer survival but this did not achieve significance. p53 protein expression showed a significant relationship to high tumour grade and a weak correlation with negative oestrogen receptor status. The data suggest that p53 protein expression may be a marker of more aggressive carcinomas but that the prognostic power of expression is likely to be weak and unlikely, therefore, to be of clinical value. The results do not resolve whether detectable p53 protein expression represents a random product of dedifferentiation, or an important feature of the malignant phenotype, playing a key role in tumour behaviour. The number of patients in our study is small, however, and investigation of a larger series is clearly indicated.     

Oestrogen and progesterone receptors in screen-detected breast carcinoma: an immunohistological study using paraffin sections

The presence of oestrogen and progesterone receptors was studied in paraffin sections of 81 screen-detected breast carcinomas using the monoclonal antibodies ER-ICA and PgR-ICA (Abbott) and the immunoperoxidase technique. The immunohistological results were compared with the results of the standard dextran-coated charcoal biochemical assay in 28 tumours which were big enough to provide tumour tissue for this assay. Sixty-three cases (78%) were oestrogen receptor positive and 62 (77%) were progesterone receptor positive. There was no statistical difference between receptor positivity in palpable or impalpable, in situ or invasive tumours. In the 28 cases where the biochemical assay was carried out, the two methods gave similar results in 23 (82%) and 21 (75%) tumours for oestrogen and progesterone receptors respectively. The majority of the remaining tumours, with one exception, were positive with immunohistology and negative with biochemistry. A good correlation was also present between the mean numerical biochemical values and the semiquantitative histological scores for both receptors. It is concluded that assessment of receptor status of small screen-detected carcinomas is feasible using routinely processed paraffin sections. There is reasonably good correlation with the results obtained by the standard dextran-coated charcoal biochemical assay, but more genuine receptor positive cases are detected by immunohistology.     

PTEN缺失型子宫内膜癌中ER、PR的表达及意义

目的：探讨ER、PR在PTEN缺失型子宫内膜癌中的表达及PTEN、ER和PR表达特征与子宫内膜癌组织临床病理特征的关系。方法采用免疫组化EnVision法检测100例子宫内膜癌组织中PTEN、ER和PR表达。结果 PTEN在子宫内膜癌患者的缺失率为52．0%(52/100);PTEN缺失型子宫内膜癌组织中ER和PR阳性率分别为15．4%(8/52)和19．2%(10/52), PTEN高表达子宫内膜癌组织中ER和PR阳性率分别为72．7%(8/11)和63．6%(7/11);子宫内膜癌患者中PTEN-ER-PR-的比率为42．0%(42/100),均明显高于其它类型(P<0．05)。不同PTEN、ER和PR表达特征与子宫内膜癌患者组织学分级和病理分期有关(P<0．05),与肌层浸润无关。结论 PTEN、ER、PR三者联合检测可能对子宫内膜癌患者预后及治疗具有重要参考价值。     

Is immunohistochemical analysis of oestrogen and progesterone receptors in endometrial carcinoma superior to the radioligand binding assay?

The aim of this study was to evaluate tissue and steroid receptor heterogeneity in endometrial carcinoma specimens as a possible source of discordance between biochemically assayed receptor status and response to endocrine treatment. For this purpose the oestrogen receptor (OR) and progesterone receptor (PR) levels in specimens from 16 endometrial carcinoma patients were analysed on adjacent tissue sections using both a radiochemical and an immunohistochemical assay. With immunohistochemical receptor analysis extensive tissue and tumour cell receptor heterogeneity was observed. Many tumour samples revealed up to 75 per cent contamination with benign tissue. In the majority of cases, evaluation of immunoreactivity in normal tissue elements of the specimen could explain the apparent discordance between semiquantitative immunohistochemical receptor scoring of tumour cells and radiochemical receptor assay. Immunohistochemical analysis of OR and PR in endometrial carcinoma specimens allows a more specific determination of tumour cell receptor content and hence may yield a more accurate prediction of response to endocrine therapy than the biochemical assay.     

子宫腺肌症雌激素受体、孕激素受体与bcl-2的表达

目的 探讨雌激素受体（ER）、孕激素受体（PR）和bcl-2在子宫腺肌症中异位和在位内膜的表达及意义。方法 用免疫组化EnVision法检测40例子宫腺肌症在位子宫内膜和肌间异位内膜ER、PR和bcl-2的表达情况。结果 在位和异位子宫内膜组织中均有ER、PR和bcl-2的阳性表达。其中腺上皮阳性表达率高于间质（P〈0．05）,且异位内膜的腺上皮bcl-2阳性表达率高于在位组织（P〈0．05）;ER和PR的表达两者差异无显著性（P〉0．05）。异位内膜腺体ER、PR与bcl-2表达具有相关性（P〈0．01）。结论 子宫腺肌症异位和在位子宫内膜组织中均有ER、PR和bcl-2的阳性表达,可能与子宫腺肌症的发生、发展有关。     

Histological grade in invasive ductal carcinoma of breast correlates with the proliferative activity evaluated by BrdU: An immunohistochemical study including correlations with p53, c-erbB-2 and estrogen receptor status

Thirty cases of invasive ductal carcinoma of the breast were classified to histological subtype according to the General Rules for Clinical and Pathological Recording of Breast Cancer of the Japanese Breast Cancer Society and histologically graded using the Nottingham method and the correlation of histology with proliferative activity was investigated using bromodeoxyuridine (BrdU). In addition, the overexpression of p53 protein, c-erbB-2 oncoprotein and estrogen receptor (ER) were immunohistochemically examined in order to discuss the relationship with histological subtype and histological grade. Histological grade correlated positively to the BrdU labeling index (LI) and overexpression of p53. High grade carcinoma demonstrated c-erbB-2 more frequently and exhibited a low incidence of ER. However, no significant relationship was found between BrdU LI, overexpression of p53 and c-erbB-2 and histological subtype. These results suggest that the histological grade does represent the proliferative activity of tumor cells and that adding the histological grade to the pathological diagnosis in invasive ductal breast carcinoma may be useful from the clinicopathological aspect concerning tumor behavior.     

Assessment of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) status in the fine needle aspirates of metastatic breast carcinomas

The assessment of estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor (HER2) status in the fine needle aspirates of metastatic breast carcinomas has prognostic and therapeutic implications. In this study, expression of ER, PR, and HER2 was assessed by immunohistochemical study in 70 cases of metastatic breast carcinomas and HER2 gene amplification was further evaluated by fluorescence in situ hybridization (FISH) in 38 (54%) cases. Positive expression of ER and PR was seen in 42 (60%) and 16 (23%) cases of metastatic breast carcinomas, respectively. HER2 immunoreactivity was scored as 0/1+ in 39 (56%), 2+ in 10 (14%), and 3+ in 21 (30%) cases. HER2 gene amplification was seen in 20% of HER2 2+ and 64% of HER2 3+ cases. ER, PR, and HER2 status in primary breast cancers were available to comparison in 31 cases (44%). The concordance rates between metastatic and primary breast carcinomas were 81% for ER, 65% for PR and 71% for HER2. Our study demonstrates that ER, PR, and HER2 status can be assessed in the fine needle aspirates of metastatic breast carcinomas and ER has a higher concordance rate between metastatic and primary breast carcinomas than PR and HER2. The addition of HER2 gene amplification FISH test helps in accurate assessment of HER2 status in metastatic breast carcinomas.     

The associations of obesity, lymph node status and prognosis in breast cancer patients: dependence on estrogen and progesterone receptor status

Breast cancer patients who are obese have a higher risk of lymph node metastases and a poorer prognosis than those who are slim. It has been claimed that estrogens derived from fat are important for these associations. If estrogens are important, these relationships must be stronger in the hormone receptor-positive than in the hormone receptor-negative groups. Body mass index (BMI) was used as a measure of obesity. The second, third, and fourth quintiles of BMI were treated as one group and termed 'medium'. Patients in the fifth quintile were termed 'obese' and those in the first quintile 'slim'. The number of women with unilateral disease treated with modified radical mastectomy and included in the study was 1211. Of all patients included, obese patients had a 1.53 higher risk of lymph node metastases compared to slim patients (p=0.02). In the PgR-negative group, obesity gave a 3.08 times higher risk of lymph node metastases (p=0.03). The risk of dying of breast cancer tended to be higher in obese than in slim patients when all patients in the study were compared (relative risk=1.38, p=0.06). BMI did not show a statistically significant relationship with prognosis if only hormone receptor status was considered. However, if lymph node status and hormone receptor status were taken together, the association was strong and reversed in the lymph node-positive group with ER-negative tumours. The adjusted relative risk was 0.33, showing that slim patients had a 3.03 (1.0/0.33) times higher risk of dying of breast cancer compared to obese patients (p=0.002). These results indicate that non-hormonal mechanisms could be important.