Immunohistochemical detection of estrogen and progesterone receptors in primary breast cancer

To evaluate the reliability of the immunohistochemical assay for estrogen receptor (ER) and progesterone receptor (PR) in the prognosis of patients with breast cancer, 83 primary tumors from the patients were studied. Immunohistochemical analysis was performed using antibody ER 1D5 for ER determination and antibody PR-ICA for PR determination. Of all tumors, ER and PR positivities were detected in 36.1% and 45.8% respectively. There was no significant relationship between ER, PR and age of the patients, tumor size or number of involved nodes. However, we found that only the immunohistochemical ER was a predictor of early recurrence in patients with primary breast cancer. In addition, there was no additive effect in recurrence-free survival when both receptor expressions were combined.     

Immunocytochemical detection of estrogen and progesterone receptors in 124 human breast cancers

Monoclonal antibodies to human estrogen receptor (ER), and rabbit progesterone receptor (PR), also recognizing human PR, were used to detect the receptors by peroxidase immunocytochemistry in frozen sections of 124 primary breast carcinomas. Both ER and PR were almost exclusively located in carcinoma cell nuclei, with heterogeneous distribution and intensity. The staining results were evaluated semiquantitatively (histoscore), based on the percentage of positively stained carcinoma cells and nuclear staining intensity. The receptor status thus determined was as follows: ER+PR+ in 50 patients, ER+PR- in 23, ER-PR- in 26, and ER-PR+ in 3 patients. There was a 79% (ER) or 70% (PR) agreement in the positivity/negativity between the immunocytochemical and steroid-binding assay (in 102 patients) with a highly significant correlation. The histoscore values increased significantly with cytosol receptor levels (ER, r = 0.623, P less than 0.001; PR, r = 0.366, P less than 0.01).     

Immunocytochemical analysis of progesterone receptors in breast cancer.

Breast cancer specimens from 116 patients were assayed for the presence of progesterone receptor (PR), with the use of a highly specific monoclonal antibody and the peroxidase-antiperoxidase technique on cryostat and permanent sections. Results were compared with those obtained by the conventional PR determination by dextran-coated charcoal (DCC) assay; they were in concordance in 90% of cryostat sections and 85% of paraffin-embedded tissue. The sensitivity and specificity of the PR immunocytochemical assay (PR-ICA) were 91% and 89% for frozen sections and 83% and 89% for permanent sections, respectively. The immunostained slides also were evaluated for several semiquantitative features, including staining intensity, heterogeneity of staining, and the proportion of positive tumor cells. A statistically significant correlation was found between the percentage of tumor cells stained with the PR immunocytochemical technique and the PR-cytosol levels (P less than 0.05). These results suggest that the PR-ICA is an effective tool in the evaluation of PR content in breast cancer and can be applied in paraffin as well as frozen sections. This technique provides excellent morphologic detail, as well as tissue localization for PR. It also offers an alternative for assessment of PR when fresh tissue is not available for conventional hormone receptor analysis. The immunocytochemical assay can be performed easily at community hospitals. Because it requires only a small amount of tissue, PR-ICA is an ideal method for analyzing specimens of insufficient size for the DCC assay. This technique also is suited to the evaluation of fine-needle aspiration biopsy specimens.     

Immunohistochemical detection of estrogen receptors in paraffin sections from primary and metastatic breast cancer

With the availability of monoclonal antibodies against the estrogen receptor (ER) it is possible to demonstrate the presence of ER immunohistochemically. Some of the antibodies are claimed to be reactive in formalin fixed, paraffin embedded tissue. We have evaluated the reactivity of one of these antibodies, D75 and found an acceptable reaction in routinely formalin fixed, paraffin embedded tissue. The antibody was applied to both primary and secondary tumors from a group of patients with recurrent breast cancer. The metastatic lesions consisted of lymph node metastases, bone marrow metastases, and liver metastases. While 41% of the primary tumors were ER-positive, this was only the case with 35%, 20%, and 17% of the lymph node, bone marrow, and liver metastases, respectively. The discordance between the ER-status of the primary tumor and the distant metastasis was 41% in cases of bone marrow metastases, and 44% in liver metastases. In most cases the shift was from an ER-positive primary tumor to an ER-negative metastasis. The results support the hypothesis that ER-negative tumor cells are probably more aggressive with a larger metastatic potential than the higher differentiated, ER-positive tumor cells.     

Immunocytochemical analysis of receptors for estrogen and progesterone in fine-needle aspirates from human breast carcinomas

Monoclonal antibodies were used for a preoperative analysis of estrogen receptors (ERs) and progesterone receptors (PgRs) in fine-needle aspirates from 44 primary human breast carcinomas. The semiquantitative receptor values obtained in cytologic specimens correlated well with those from enzyme immunoassay analysis on surgically removed tumor tissue (r = 0.74 for PgR; r = 0.75 for ER). Cytologic smears showed a heterogenous tumor cell distribution of ER and PgR in 29 and 23 cases, respectively. The results suggest that measurement of the ER and PgR in cytologic smears is an accurate and reliable technique that can be performed on a minimum amount of tissue.     

The effects of glyoxal fixation on the histological evaluation of breast specimens

Glyoxal (GL), a non-formalin-containing aldehyde tissue fixative, is advocated as a superior fixative that is environmentally safe and lacks the purported carcinogenic health hazards associated with formalin use. In addition, it is advertised as requiring no antigen retrieval before immunohistochemical staining. We compared GL fixation to standard formalin fixation of breast specimens removed for microcalcifications or breast tumors. Although the hematoxylin and eosin morphology of GL-fixed and formalin-fixed tissues was equivalent, detection of microcalcifications in GL-fixed breast specimens was hampered by loss of basophilia, likely due to increased calcium solubility in glyoxal. Moreover, estrogen receptor detection in GL-fixed specimens was diminished compared to formalin and did require antigen retrieval.     

Comparative evaluation of ER-ICA and enzyme immunoassay for the quantitation of estrogen receptors in breast cancer

Evaluation of estrogen receptors (ERs) is essential for the treatment and prognosis of human breast cancer. The authors have undertaken a study of 225 primary breast cancers to compare the receptor values using immunohistochemistry and enzyme immunoassay (EIA) techniques. The results are compared with the biochemical assay with the use of the dextran-coated charcoal (DCC) method done on the same specimens. The overall concordance between ER-ICA and DCC was 77%. Ninety-seven percent concordance was observed for specimens with a positive ER-ICA score (greater than 70 and the DCC more than 10 fmol/mg protein). Ninety-two percent of tumors had negative results by ER-ICA when the DCC was less than 10 fmol/mg protein. When the ER-ICA score was plotted against the DCC or EIA value, a significant correlation was obtained with a P value of less than 0.001. The correlation between the ER-ICA score and the total ER content (cytosol and nuclear) assayed by EIA was similar to comparison with cytosol alone. The authors conclude that the ER-ICA can be useful in assessing ERs in breast especially in hypocellular or small tumors and in assessing the degree of heterogeneity in a given tumor. The immunohistochemical and biochemical assay methods are complementary and provide greatly needed information for the management of breast cancer.     

新辅助化疗乳腺癌病理标本的处理对策及报告规范

新辅助化疗(neoadjuvant clhemotherapy)是指在进行手术治疗前对肿瘤患者进行化疗的一种治疗方法.1973年,米兰癌症中心的研究者首先采用新辅助化疗对失去手术机会的乳腺癌患者进行治疗,以期缩小肿块从而使患者有机会接受手术或放射治疗 [1].与常规化疗相比,新辅助化疗具有以下一些特点:(1)约80%的患者化疗后肿块缩小,原本失去手术机会的乳腺癌患者可重获手术机会;一些原本需施行乳房全切除的患者可进行保乳治疗,虽然这些患者的局部复发率可能升高,但其存活率与先手术后化疗的患者相当.     

Immunocytochemical assay of progesterone receptors in paraffin-embedded specimens of endometrial carcinoma and hyperplasia: a preliminary evaluation

An immunohistochemical method for the detection of progesterone receptors (PgR) using the monoclonal anti-PgR antibody KD 68 was utilized to study paraffin-embedded tissue sections from women with endometrial carcinoma and hyperplasia. Stromal as well as myometrial nuclear PgR were nearly always apparent. In carcinoma, 11/24 (46%) of cases showed epithelial positivity, whereas in hyperplasia 8/9 (89%) were PgR-positive (P less than 0.05). Initial biochemical PgR assays by the dextran-coated charcoal method were compared with results of PgR-immunocytochemical assays (ICA) in the paraffin-embedded tissue and were in concordance in 92%. In the one discordant specimen, PgR-ICA-negative tumor cells were seen infiltrating PgR-ICA-positive myometrium, and the biochemical assay was thus felt to be falsely positive. Twelve additional cases of endometrial carcinoma were also studied for estrogen receptor (ER) by immunocytochemistry. Two were positive for both ER and PgR, while five were negative for both receptors. The immunocytochemical methods described allow for analysis of routinely fixed, paraffin-embedded specimens, thus permitting analysis of very small specimens and archival material.     

A comparative immunohistochemical and biochemical study of estrogen receptors in breast cancer

The authors review the results of a comparative study of 25 mammary carcinomas, examined by the immunohistochemical method (IHM) with the use of ER-D5 monoclonal antibodies (Amersham) and by the cytosol receptor protein binding of a labeled ligand followed by the unbound hormone adsorption on dexteran-coated coal (DCM). Pap test with the above antibodies has been employed in immunohistochemical studies. The IHM has been highly specific. The results coincided in 21 of the 25 cases (83.3%). The reaction has been heterogenous in the same tumor. Quantitative values obtained by the two methods coincided in many cases. The IHM may have a wide practical application, provided the reagents are available, for it does not require sophisticated equipment and is simple to perform.     

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.     

Immunocytochemical evaluation of HER-2/neu on fine-needle aspirates from primary breast carcinomas

Detection of HER-2/neu alterations is increasingly used in breast cancer patients for therapeutic purposes. This study examines the reliability of HER-2/neu immunocytochemical assessment on 66 cytospin smears obtained by fine-needle aspiration biopsy from breast cancer patients. Results were compared with those obtained by both fluorescence in situ hybridization (FISH) on fine-needle aspirate (FNA) and immunohistochemistry (IHC) on matched histologic section. Concordance between immunocytochemistry (ICC) and FISH was 78% and between ICC and IHC was 84%. Discordance mainly concerned seven unamplified cases that resulted positive by ICC and four cases scored negative by IHC but positive by ICC. Simultaneous assessment of HER-2/neu by ICC, IHC, and FISH was available in 24 cases; the concordance was 75%. In this study, the false positivity of immunocytochemical technique represents the major criticism. In our experience, FISH remains the most objective and powerful technique for HER-2/neu assessment on breast cancer FNAs.     

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.     

Immunocytochemical characterization of male breast cancer.

Biologic properties of breast cancer in men that might reflect alterations in pathogenesis from the disease in women were examined. We studied 22 tumors from males, 18 invasive carcinomas, three of which were papillary, and three in situ tumors of which one was papillary, and one papilloma. Our data support the previously reported high incidence of papillary carcinoma in men. Estrogen receptor status and the expression of cancer-associated antigens recognized by antibodies DF3, B73.2, SP-1, and c-erbB-2 were compared to matched tumors from females. Immunocytochemistry was performed on formalin-fixed, paraffin-embedded sections using standard avidin-biotin techniques; anti-PSA was used to exclude the possibility of metatastic prostate cancer, and 12 cases of gynecomastia were included as nonmalignant controls. The incidence of estrogen receptor positivity was higher in tumors from males (73%) than from females (54%), as has been reported previously. The range of expression of all breast cancer antigens tested in male tumors was similar to that observed in females, but some interesting differences were noted. With the exception of the anti-mucin DF3, all the antibodies reacted only with neoplastic tissues. Expression of the oncoprotein c-erbB-2 was lower (17%) in males than in females (33%), despite the preponderance in men of the large-cell type carcinomas that have been associated with c-erbB-2 expression. Unexpectedly, the pregnancy-associated hormone detected by SP-1 was expressed in 33% of tumors from males and, in contrast to females, was found in less differentiated tumors.     

抗原微波热修复对乳腺癌雌激素受体和孕激素受体免疫组织化学检测结果的影响

免疫组织化学是目前乳腺癌激素受体检测的常规方法,但免疫组织化学检测易受多种因素的影响,其标准化程度令人担忧.英国国家免疫细胞化学外部质量评估方案(NEQAS-ICC)曾评估200余个实验室对同一例乳腺癌组织进行的雌激素受体(ER)染色,发现仅36%的实验室获得准确的检测结果[1],提示抗原修复是影响免疫组织化学检测结果最主要的因素,而激素受体水平低的癌组织进行免疫组织化学染色更易受到各种因素的影响.     

Expression of estrogen and progesterone receptors and HER-2 by breast cancer cells

Breast cell carcinomas removed from 62 women aged from 33 to 74 years were studied immunohistochemically. 24 tumours were of T1N0M0, 5-T1N1M0, 20-T2N0M0, 11-T2N1M0 and 2-T2N0M0. Invasive ductal carcinoma occurred in 59.6%, invasive ductal in 40.4%. Estrogen receptor expression was found in 59.6%, progesterone in 38.7%. Simultaneous expression of these receptors was in 35.5%. Estrogen receptor and progesterone receptor expression were registered in 24.2 and 3.2% cases, respectively. There was neither estrogen nor progesterone expression in 37%. HER-2 expression was observed in 45.2%, the highest being at the age from 50 to 55 years and in invasive ductal carcinoma. Enhancement of estrogen receptor expression was accompanied with lower rate of HER-2 expression. All three antigens expression does not depend on the presence or absence of metastases to the regional lymph nodes.     

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.     

Immunocytochemical evaluation of estrogen receptor on archival Papanicolaou-stained fine-needle aspirate smears

The availability of limited fine-needle aspirate smears necessitates the selection of immunocytochemical (IC) methods that allow reuse of Pap-stained smears to assess the estrogen receptor (ER) status of breast carcinoma. The objective of the current study was to compare IC evaluation of ER status on FNA smears by three methods: 1) ER-ICA using H222 monoclonal antibody performed on slides fixed in formaldehyde-methanol-acetone; 2) destained Pap slides using 1D5 antibody; and 3) Pap-stained slides without destaining using the same 1D5 antibody. Two representative Pap smears of breast carcinoma were selected from 48 cases of breast carcinoma in which ER was previously evaluated by ER-ICA. One of these Pap smears was used as such and the other was destained prior to immunostaining by a modified ABC method using 1D5 monoclonal antibody. The number of cells with positive nuclear staining was expressed as a percentage and the intensity of staining was semiquantitatively scored on a scale of 1+ to 3+. The degree of agreement between the three methods was evaluated statistically by weighted kappa statistics. Thirty cases (63%) showed varying degrees of positive staining while 18 cases (38%) were entirely negative by all three methods. Significant discrepancies in the number of cells with positive staining and in the intensity of staining between the three methods occurred in 40% and 23% of the cases and was mainly due to a reduction in the number of cells with positive staining and the intensity of staining using Pap slides in comparison to ER-ICA. Weighted kappa agreement of the percentage of cells with positive staining using Pap-stained slides and destained Pap-slides in comparison to ER-ICA was 0.75 and 0.64, respectively, and that for the intensity of staining was 0.75 and 0.66, respectively. Therefore, IC evaluation of ER using Pap-stained smears as such or destained Pap smears compared favorably with ER-ICA. However, Pap-stained smears used as such for ER immunostaining showed a slightly better agreement with ER-ICA than destained Pap smears. Because significant differences in ER-IC staining can occur with any of the immunocytochemical methods, a negative result is less reliable as an indicator of true ER status than a positive result.