Estrogen receptor (ER), progesterone receptor (PR), and HER2 expression pre- and post- neoadjuvant chemotherapy in primary breast carcinoma: a single institutional experience

Background

The estrogen receptor (ER), progesterone receptor (PR), and HER2 profile of a primary breast carcinoma plays a significant role in patient management and treatment. Because of the increasing utilization of neoadjuvant chemotherapy or hormone therapy, surgically-resected carcinomas often show marked treatment effect. The aim of this study was to compare immunohistochemical (IHC) profiles (ER, PR, HER2, HER2 FISH) of primary breast carcinomas before and after neoadjuvant chemotherapy to assess the subsequent effects on hormone receptor status.

Design

Primary breast carcinomas from 38 female patients treated with neoadjuvant therapy after needle core biopsy or fine needle aspiration diagnosis were included. Histologic data was collected for each case, including site, type, grade, tumor size (cm), pre- and post- neoadjuvant treatment IHC panel (ER, PR, HER2), and fluorescence in-situ hybridization (FISH) for HER2.

Results

Of the 38 carcinomas studied, 45 % were positive for ER by IHC both pre- and post- neoadjuvant treatment (P=1.00). IHC studies for PR in these 38 patients showed 37% positivity for PR pre-neoadjuvant therapy and 21% positivity post-treatment (p=0.03). For 37 patients with HER2 IHC, 32% were positive pre-treatment, and 22% were positive post-treatment (P = 0.20). For 7 patients, HER2 FISH was positive in 71% pre-therapy and in 57% post-treatment (P=0.32).

Conclusions

Profiles for ER, HER2 IHC, and HER2 FISH were not significantly different in primary breast carcinomas before and after neoadjuvant chemotherapy. Further investigation is warranted to assess reproducibility of technique and investigate clinical implications of significant loss of PR status in treated patients.     

Human epidermal growth factor receptor 2 expression in breast cancer: correlation with clinical pathological features

The overexpressed HER2 (human epidermal growth factor receptor 2) is a valuable therapeutic target. Precise assessment of HER2 status is thus crucial in the treatment of breast cancer. In this study, formalin-fixed, paraffin-embedded samples of tumors from 304 breast cancer patients who underwent curative surgery procedures between 2011 and 2014 were tested by immunohistochemistry (IHC) as a primary estimate of HER2 status, followed by fluorescence in situ hybridization (FISH). Concordance rate between IHC and FISH was evaluated. The Χ² test was used to evaluate the correlation between HER2 gene amplification status and different clinical pathological features including: (estrogen receptor) ER and (progesterone receptor) PR expression, age, menopausal status and tumor size. The results show that 84.8% of IHC score 3+ cases and 6.2% of IHC score 0/1+ cases were amplified by FISH. After exclusion of group IHC 2+, the concordance rate between FISH and IHC was 87.4%. There was a significant inverse association between expression of hormone receptors (ER and PR) and HER2 amplification (P < 0.001) among the patients studied. However, no relationship was observed between HER2 amplification and age, menopausal status and tumor size (P > 0.05). The data demonstrate a relatively high level of concordance rate for HER2 testing between FISH and IHC, and HER2 overexpression was associated with the levels of ER and PR.     

Estimation of hormone receptor status and HER2 in cytologic cell blocks from breast cancer using the novel rabbit monoclonal antibodies (SP1, SP2, and SP3)

The determination of estrogen (ER) and progesterone (PR) receptor status has become standard practice in the evaluation of patients with invasive breast cancer, having important prognostic and therapeutic implications. HER2 assessment is important to evaluate the response to Herceptin® (Trastuzumab) therapy for primary and metastatic breast cancer. This study is undertaken to compare rabbit monoclonal antibodies (RabMAb) for ER, PR, and HER2 against FDA‐approved monoclonal and polyclonal antibodies (FDAMpab). Cell blocks from primary and metastatic/recurrent breast carcinomas of 52 breast cancer patients were used. Immunohistochemistry was performed, following optimized epitope retrieval, with a polymer based detection system using RabMAb: ER (SP1), PR (SP2), and HER2 (SP3). FDA approved Mpab (Dako) used were: ER (1D5); PR (PgR636); and HercepTest® kit according to manufacturer's instructions. HER2 immunostain is correlated with FISH results. Overall, positive, and negative agreement is as follows: 88.5, 88.9, and 88.2% for ER; 84.6, 70.5, and 91.4% for PR; 58.3, 100, and 50% for HER2. There is substantial to moderate agreement between RabMAb and FDAMpab for ER (κ = 0.75) and PR (κ = 0.64), respectively. There is poor agreement (κ = 0.25) between RabMAb (SP3) and FDApab (HercepTest®). SP3 shows better concordance (93.8%) than HercepTest® (46.9%) with FISH results. RabMAb SP clones are almost comparable with FDA‐approved ER and PR, with fair to moderate agreement. Both are as sensitive as their FDA‐approved clones. SP3, on the other hand, is superior to HercepTest® for detecting HER2 overexpression, with an excellent concordance with FISH. Diagn. Cytopathol. 2009. © 2009 Wiley‐Liss, Inc.     

Oestrogen receptor/progesterone receptor and human epidermal growth factor receptor 2 status in breast cancer: a 9-year study at Princess Noorah Oncology Center, Saudi Arabia

Satti M B
(2011) Histopathology 59 , 537–542 Oestrogen receptor/progesterone receptor and human epidermal growth factor receptor 2 status in breast cancer: a 9‐year study at Princess Noorah Oncology Center, Saudi Arabia Objective: To determine the oestrogen receptor/progesterone receptor (ER/PR) and human epidermal growth factor receptor 2 (HER2) status in Saudi Arabian patients presenting with breast cancer to Princess Noorah Oncology Center (PNOC) and to explore the correlation of these markers to each other, to tumour type and to grade. Methods and results: Pathology material and records of symptomatic patients presenting to the centre during 2001–2009 were reviewed for patients’ age, tumour size, type and grade and ER/PR and HER2 immunohistochemistry (IHC) status using the Dako HercepTest Kit as well as fluorescence in situ hybridization (FISH) for HER2 IHC 2+ score cases, as per the 2007 American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) guidelines. There were 852 cases, with a mean age of 49 years and a mean tumour size of 3.0 cm with 75% node positivity. Of all cases, 772 (90.6%) were ductal carcinoma; 64% were ER/PR⁺ and 23% were HER2⁺; triple‐negative cases accounted for 24%. Conclusions: ER/PR and HER2 status did not differ from that reported previously, showing a direct correlation to tumour type and grade of ductal carcinoma. However, a difference exists in the relatively lower ER positivity in patients aged >50 years and the higher percentage of triple‐negative cases. This study would serve as a baseline for other future national studies and for planning strategies to targeted therapy.     

Absence of estrogen receptor alpha (ESR1) gene amplification in a series of breast cancers in Taiwan

Immunohistochemical expression of ERα, encoded by the ESR1 (estrogen receptor 1) gene located at 6q25.1, is the most important determinant of responsiveness to endocrine therapy in breast cancer. The prevalence and significance of ESR1 amplification in breast cancer remain controversial. We set out to assess ESR1 status and its relevance in breast cancer in Taiwan. We tested tissue samples from 311 invasive carcinomas in a tissue microarray for ESR1 status by fluorescent in situ hybridization (FISH) and chromogenic in situ hybridization (CISH). In order to examine its association with ERα and ESR1 status, HER2 status was determined by FISH. Of the carcinomas, 58.8 % (183/311) was ERα positive. None of the carcinomas showed amplification of ESR1 by either method, whereas 24.1 % (75/311) of the carcinomas harbored HER2 amplification. Of the carcinomas, 9.6 % (26/301) showed ESR1 gain (1.3?≤?ratio ESR1/chromosome 6?<?2) by FISH and 10 % (24/299) by CISH. FISH and CISH results showed a good correlation (κ-coefficient?=?0.786). ESR1 gain by FISH and CISH was significantly associated with high-grade (P?=?0.0294 and 0.0417, respectively) but not with ERα expression, HER2 status, or overall survival. ERα positivity was significantly associated with better overall survival (P?=?0.039). HER2 amplification was significantly related with poor overall survival (P?=?0.002). Our data confirm that in breast cancer, HER2 amplification is a frequent genetic aberration and a negative prognostic factor, and show that ESR1 amplification is not a key genetic abnormality in the tumorigenesis of breast cancer in Taiwan.     

Chromogenic in situ hybridisation testing for HER2 gene amplification in breast cancer produces highly reproducible results concordant with fluorescence in situ hybridisation and immunohistochemistry

AIMS: The objective of this study was to evaluate the accuracy, ease of use and reproducibility of chromogenic in situ hybridisation (CISH) for HER2 testing by studying its inter-laboratory concordance in five Australian pathology laboratories. METHODS: The HER2 status of 49 breast cancers was determined by CISH twice in two different laboratories. Each sample had previously been tested by immunohistochemistry (IHC; 2+ and 3+ cases selected) and fluorescence in situ hybridisation (FISH). Participating laboratories were blinded to these test results. Oestrogen receptor (ER) status was also evaluated for each cancer. RESULTS: High correlation was observed between FISH and CISH results. No cases showing high gene amplification by FISH were scored as non-amplified by CISH (kappa coefficient = 1). High correlation was observed between IHC and CISH, all IHC 3+ samples showing amplification by CISH. Inter-laboratory CISH concordance was also good (kappa coefficient = 0.67). Fifty-six per cent of HER2-amplified samples tested ER positive, while 42% of ER-positive cases showed HER2 gene amplification, confirming that HER2 testing should not be confined to ER-negative breast cancers. CONCLUSIONS: These findings demonstrate that CISH is a robust test to assess HER2 status in breast cancer and therefore is an important addition to the HER2 testing algorithm.     

Discordance in receptor status between primary and metastatic breast cancer and overall survival: A single-center analysis

BackgroundThe tumor phenotype may change between primary and metastatic breast cancer. We compared the expression of estrogen receptor (ER), progesterone receptor (PR), and HER2 in a series of primary breast carcinomas (PBC) with their metastatic relapses and analyzed the impact of any changes on survival.Materials and methodsIt was a single-center retrospective study, collecting consecutive cases of metastatic breast carcinoma diagnosed in the pathology and medical oncology departments at Habib Bourguiba University Hospital in Sfax, Tunisia. An immunohistochemical study was used to assess ER, PR, and HER2 expression. Overall survival (OS) and post-metastasis survival (PMS) were evaluated using multivariable Cox regression analysis.ResultsOur study included 68 patients. ER and PR status changed in 29.4 % and 39.7 % of cases, respectively. Conversions were mainly from positive to negative status (22 % and 23.5 % for ER and PR, respectively). Differences in HER2 status were observed in 19.6 % of cases, with loss of overexpression in 6 patients (10.7 %). Adjuvant trastuzumab therapy and PBC molecular subtype (HR-, HER2+) were associated with HER2 status discordance (p = 0.02 and 0.03, respectively). On multivariable analysis, HR-negative conversion tumors were significantly associated with a worse OS (p = 0.042) and PMS (p < 0.001), compared to HR-concordant positive tumors.ConclusionThis study establishes that HR and HER2 status discordance between primary and metastatic breast carcinoma has a prognostic impact on patient outcome. Analyzing these receptors' status in all newly diagnosed cases of metastatic breast carcinoma is strongly recommended and would provide information for changing treatment strategies.     

Simultaneous detection of HER2/neu gene amplification and protein overexpression in paraffin-embedded breast cancer

The determination of HER2/neu status in breast carcinomas has become essential for the selection of breast cancer patients for Herceptin therapy. Herceptin treatment is used in patients with metastatic breast carcinoma with HER2/neu protein overexpression detected by immunohistochemistry (IHC) or gene amplification analysed by fluorescence in situ hybridization (FISH). A multiparametric fluorescent approach based on the simultaneous detection of HER2/neu gene amplification and protein expression was established to increase the accuracy, and to improve the reproducibility, of HER2/neu diagnostics. Based on four paraffin-embedded breast cancer cell lines, a combined fluorescent immunostaining (FIHC) and FISH method was developed by using the PathVysion HER2 DNA Probe Kit (VYSIS) and the polyclonal antibody from the HercepTest (DAKO). Diagnostic applicability was documented on 215 formalin-fixed primary breast carcinomas. Criteria for immunofluorescence quantification were chosen by analogy with the FDA-approved HercepTest scoring, ranging from 0 to 3+. There was 97.7% concordance between conventional IHC and fluorescence IHC. The FISH data resulting from the multiparametric approach did not differ from conventional FISH. Breast carcinomas with HER2/neu protein overexpression and simultaneous gene amplification were detected with 100% sensitivity. In addition, five of the 215 cases (2.3%) had HER2/neu gene amplification without protein overexpression. The main advantage of this novel approach is that polysomy, aneuploidy, gene amplification, and protein content can be analysed simultaneously in the same cell.     

Reliability of histological malignancy grade,ER and HER2 status on core needle biopsy vs surgical specimen in breast cancer

Early determination of malignancy grade and biomarkers in primary breast cancer is important when planning the treatment with neoadjuvant therapy and breast‐conserving surgery. The aim of this study was to assess the reliability of malignancy grade on core needle biopsies, and to compare the concordance of ER and HER2 status on core needle biopsies vs surgical specimens. Eighty‐nine patients with invasive ductal or lobular carcinoma were included. Malignancy grade was upgraded by one degree in 23.0%. The mitotic count had most impact on the malignancy grade, and may be underestimated on core needle biopsies compared with surgical specimens. ER status was concordant in core needle biopsies and surgical specimens in 98.0%, but should be repeated when doubtfully negative. HER2 status was concordant in 84.0% prior to FISH‐HER2. Including FISH‐HER2 data, the core needle biopsies showed a final total concordance of 95.4%. In conclusion, ER and HER2 are useful markers in core needle biopsies, providing a reliable base for preoperative decisions in neoadjuvant chemotherapy, and may be omitted on surgical specimens with certain precautions and exceptions. Determination of malignancy grade should be repeated on surgical specimens, and one should be aware of underestimation on core needle biopsies.     

Validity of immunohistochemistry method in predicting HER‐2 gene status and association of clinicopathological variables with it in invasive breast cancer patients

Human epidermal growth factor receptor‐2 is an important and prognostic factors and one of the most targeted proteins in breast cancer's therapy. There is no globally accepted method for determining its status. Here, we aimed to evaluate the immunohistochemistry method validity in predicting HER‐2 status by Fluorescence in situ hybridization method and investigate some clinicopathological variables association with HER‐2 amplification. A total of 190 HER‐2 2+ and 3+ by immunohistochemistry (IHC) invasive breast cancer cases were enrolled in this study. Fluorescence in situ hybridization (FISH) was performed for these cases using FDA criteria and the association between clinicopathological variables and HER‐2 status evaluated. Study consisted of 190 invasive breast cancer patients (160 HER‐2 2+ and 30 HER‐2 3+). HER‐2 FISH amplification according to FDA criteria was found 27.5% (44/160 patients) in HER‐2 2+ patients and 83.3% (25/30 patients) in HER‐2 3+ patients. Tumors with HER‐2 amplification were more likely to be ER‐negative (51.0% vs 31.2%, p = 0.013) and PR‐negative (52.9% vs 27.0%, p < 0.001). This study showed that immunohistochemistry is not a good method for evaluating HER‐2 status and decision‐making about trastuzumab therapy even with 3+ score patients. However, this result may not be too strong for IHC 3+ cases due to the limited number of these patients in this study.     

HER2 FISH analysis on a skeletal metastasis: a case report and technical review

Eighty per cent of patients who develop metastatic breast cancer will experience a skeletal metastasis. National guidelines recommend biopsy and repeat estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor-2 (HER2) testing for all metastatic breast cancers to guide further treatment planning. Biomarker analysis in bone biopsy samples is challenging due to the necessity of decalcification of the bone sample in order to allow tissue sectioning. The decalcification solutions may result in the degradation of proteins and thus affect the results of immunohistochemical (IHC) testing for ER, PR, and HER2. The decalcification solutions may also degrade nucleic acids, and result in poor or no hybridization signals when attempting in situ hybridization (ISH) testing for HER2. Here, we report a patient with metastatic breast cancer to the bone who underwent image guided core needle biopsy. Her past medical history was significant for ER+, PR+, and HER2+ breast cancer. The biopsy yielded cores of bone, which were decalcified, and a blood clot that was separated from the bone and fixed in formalin without decalcification. Both specimens showed metastatic carcinoma consistent with a breast primary when viewing hematoxylin and eosin (H&;E) stained sections. Immunohistochemical analysis, which was performed only on the clot sections, showed ER+, PR???, and HER2 equivocal (2+) tumor cells. Immunohistochemical analysis was not performed on the bony tissue, because our assays are not validated for use on decalcified tissue. Human epidermal growth factor receptor-2 fluorescence in situ hybridization (FISH) analysis was attempted on the bone biopsy and yielded no signals (analytic failure). This is typical of HER2 FISH analysis in decalcified specimens, due to the impaired deoxyribonucleic acid (DNA) quality. However, HER2 FISH analysis on the clot sections showed that the HER2 gene was amplified. Given the clinical importance of repeat HER2 testing in patients with recurrent breast cancer, techniques that avoid decalcification are needed. Separating the blood clot from the bony tissue, and processing it without decalcification may yield viable tumor whose DNA has not been degraded, and it then can be used for the analysis of ER, PR, and HER2.     

Comparison of the expression of prognostic biomarkers between primary tumor and axillary lymph node metastases in breast cancer

The prognosis and prediction of axillary lymph node (ALN) metastases in breast cancer is traditionally based upon the biomarkers status of the primary tumor. Some retrospective studies showed significant discordance in receptor expression between primary and metastatic tumors. We aim to prospectively assess the incidence of discordant biomarkers status in primary tumor and ALN metastases and to evaluate the role of ALN biopsies for the reassessment of receptor status. Tissue arrays were constructed from 54 breast cancer patients with ALN metastases diagnosed. Arrays were immuno-stained to compare protein expression of four biomarkers including estrogen receptor (ER), progesterone receptor (PR), HER2, and Ki67 by immunohistochemistry. The kappa value of consistency in the primary tumor and the metastatic lymph nodes were 0.465 for ER, 0.445 for PR, and 0.706 for HER2. Good consistency was shown for Ki67 expression in primary and metastases regions with T test. No significant difference is existed between primary tumor and ALN metastases. It is concluded that the good consistency is present for ER, PR, HER2 and Ki67 between the primary tumor and the metastatic lymph nodes, suggesting that ER, PR, HER2, or Ki67 status in primary tumors could reflect their status in ALN metastases.     

显色原位杂交和免疫组织化学检测乳腺癌HER2/neu基因状况和蛋白表达的对照性研究

目的探讨显色原位杂交（CISH）在检测乳腺癌中HER2／neu基因扩增上的作用。方法挑选乳腺浸润性导管癌患者组织石蜡蜡块（回顾性255例,前瞻性271例）,进行免疫组织化学（IHC）、CISH检测。15例回顾性标本送往德国HERA检测中心进行FISH检测。结果（1）回顾性病例中IHC阳性3＋者CISH基因扩增率为91．6％（120／131）,IHC2＋者CISH基因扩增率为56．5％（39／69）,IHC与CISH检测结果符合率为81．2％（207／255）,两者明显相关（P〈0．01）。（2）前瞻性病例中IHC蛋白过表达率31．7％．CISH基因扩增率27．3％。IHC3＋者CISH基因扩增率为91．4％（53／58）,IHC2＋者CISH基因扩增率为46．4％（13／28）,IHC与CISH检测结果符合率为89．7％（243／271）,两者明显相关（P〈0．01）。（3）经德国检测中心荧光原位杂交（FISH）检测的15例中14例和CISH结果完全一致,1例检测失败,而CISH为无扩增。（4）CISH检测基因扩增与雌激素受体（ER）、孕激素受体（PR）表达明显负相关（P值均〈0．01）。结论CISH检测HER2基因扩增结果与IHC检测蛋白过表达及FISH结果高度一致,CISH是检测HER2基因扩增的一项新技术。     

Feasibility of using tissue microarrays for the assessment of HER-2 gene amplification by fluorescence in situ hybridization in breast carcinoma.

Tissue microarrays (TMAs) have been commonly used to study protein expression by immunohistochemistry (IHC). However, limited data exist on the validity of using TMAs to study gene amplification. In this study, we evaluated the feasibility of using breast carcinoma TMAs to study HER-2 gene amplification by fluorescence in situ hybridization (FISH). In addition, hormonal receptor status (ER and PR) and HER-2 protein overexpression by IHC were also studied, and results were compared with whole tissue sections. FISH for HER-2 was performed on formalin-fixed paraffin-embedded tissue from 114 invasive breast carcinomas both on whole tissue sections and on TMAs containing the same tumors. The TMA was created using 0.6-mm tissue cores with four sampled cores per tumor from the same tissue block used for whole section FISH. The PathVysion HER-2 probe kit was used for the FISH analysis. A ratio of HER-2:Chromosome17 > or =2.0 was interpreted as positive for gene amplification. The ER or PR was interpreted as positive when nuclear staining was detected in more than 10% of tumor cells. The HER-2 IHC (HercepTest; DAKO Corp, Carpinteria, CA) results were interpreted as 0, 1+, 2+, and 3+ according to standard criteria. The FISH results in the TMA and whole sections were concordant in 99 out of 101 successfully analyzed cases (99%). The FISH scores were consistent among the two to four cores in the majority of the cases. ER and PR results were concordant between whole sections and TMA cores in 97% (107/110) and 89% (97/109) cases, respectively. The overall concordance for HER-2 status by IHC between whole sections and TMA cores was 86% (94 out of 109 cases). TMAs are a reliable approach to study HER-2 gene amplification in a high throughput manner.     

Tissue microarrays for routine diagnostic assessment of HER2 status in breast carcinoma.

The use of tissue microarray (TMA) technology may substantially reduce the costs of routine testing of breast carcinomas for human epidermal growth factor receptor 2 (HER2) status. After a preliminary pilot study comparing the TMA results with those obtained on whole section, which showed an excellent agreement (with kappa values >0.90) for both immunohistochemical and fluorescent in situ hybridization (FISH) method, we introduced the TMA technique in our routine work. A total of 1158 invasive breast carcinomas were submitted for the determination of HER2 status, which was assessed in 74 weekly runs. One hundred twenty-five of 1084 surgical specimens (11.5%) were judged as unsuitable for inclusion into TMAs. In 32 of 959 tumors included in TMAs (3.3%), the respective cores were uninformative, and HER2 status was determined on whole sections. Thus, HER2 status was finally determined on TMA in 927 cases (81.1%). A typical weekly run comprised 1 TMA (consisting, on average, of 13 tumors), 2 whole sections of surgical specimens and 1 whole section of core needle biopsy, and the number of processed slides for each method decreased from 16 to 4 per week. In all, 14.7% of tumors were HER2 positive by FISH. In both TMAs and whole sections, immunohistochemical results were in good agreement with FISH for cases scored as 0/1+ (98% and 97%) and for those scored as 3+ (96% and 87%), whereas concordance was poor in cases scored as 2+ (30% and 13%, respectively).     

Immunohistochemical detection of estrogen receptor,progesterone receptor and human epidermal growth factor receptor 2 in formalin‐fixed breast carcinoma cell block preparations: Correlation of results to corresponding tissue block (needle core and excision) samples

Evaluation of ER, PR and Her 2 are routinely performed on breast carcinomas. For accurate detection of these markers, compliance with the ASCO/CAP guidelines is recommended. Our previous study showed that alcohol fixation did not affect ER results when alcohol‐fixed cell block (CB) sections were compared to formalin‐fixed tissue sections, while PR and Her2 showed less concordance. The aim of this study was to evaluate and to compare ER, PR and Her2 IHC results on formalin‐fixed CB sections to those observed on subsequent surgical (needle core or resection) specimens (SS). Fifty cases of formalin fixed CB samples obtained from primary (18%) and metastatic (82%) breast carcinomas were studied, all of which had subsequent SS available. ER, PR, and Her2 IHC studies were done on all samples and results were compared. ER results on formalin‐fixed CB samples showed excellent correlation with SS (correlation coefficient cc = 0.82). While there was minimal improvement in PR results (cc = 0.433), Her2 detection did not improve by formalin fixation (cc = 0.439). Formalin fixation for CB preparations does not significantly improve the already good detection of ER positive breast tumors. The concordance rate in PR and IHC results between formalin‐fixed CB and SS samples showed improvement as compared with the alcohol‐fixed CB results. However, there was no improvement in detection of Her2 overexpression by using formalin fixation on cytology specimens. Diagn. Cytopathol. 2013. © 2012 Wiley Periodicals, Inc.     

Hormone receptor status and HER2/neu overexpression determined by automated immunostainer on routinely fixed cytologic specimens from breast carcinoma: correlation with histologic sections determinations and diagnostic pitfalls

Although fine-needle aspiration cytology is a routine procedure for the diagnosis of breast carcinoma, cytologic specimens have rarely been used for evaluation of hormone receptor status and HER2/neu overexpression. In order to compare the biological markers on cytology and on histology, routinely fixed smears of 110 primary breast carcinomas were immunostained for estrogen receptor (ER), progesterone receptor (PgR), and HER2/neu by automated immunostainer and the results were compared with the corresponding histologic sections. ER was expressed in 76 of 110 (69%) cases and PgR was expressed in 51 of 110 (46%). Overexpression of HER2/neu was observed in 30 of 110 (27%) cases. Concordance between cytology and histology was 98% for ER, 95% for PgR, and 100% for HER2/neu. There was no false positive result on smears. Diagnostic pitfalls in determination of hormone receptor status on smears included intratumoral heterogeneity and presence of mucin.     

Evaluation of new monoclonal antibodies in detection of estrogen receptor,progesterone receptor,and Her2 protein expression in breast carcinoma cell block sections using conventional microscopy and quantitative image analysis

Accurate assessment of estrogen receptor (ER), progesterone receptor (PR), and Her2 status of breast carcinomas is critical for predicting response to systemic therapies. Recently, developed rabbit monoclonal antibodies (RMab) are reported to have higher sensitivity than murine monoclonal antibodies (Mab). This study compares RMabs against FDA‐approved Mab (FMab) in breast carcinoma cell block sections using visual and image quantification. Cell blocks from 52 breast cancers were studied. Immunohistochemistry using RMab (ER, PR, and Her2) was compared with FMabs (ER, PR, Dako) and HercepTest (HerFDA). Fluorescent in situ hybridization (FISH) was used as a reference standard for Her2. Slides were later scanned and reanalyzed with an automated cellular imaging system (ACIS III, Dako). Frequency of ER (38.5% vs. 36.5% for visual; 55.8% vs. 57.7% for image) and PR (28.8% vs. 36.5% for visual; 50% vs. 51.9% for image), and concordance (overall agreement is 71.2% and 75% for visual and image ER; and 84.6% and 59.6% for visual and image PR) were similar for both FMab and RMab, respectively. Overall agreement (53.8% vs. 77.1% for visual and image detection, respectively, using HerFDA and RMab) is poor to moderate for Her2. Visual Her2 (RMab) has the highest concordance (94.1%), and visual HerFDA has the lowest concordance (35.3%) with FISH. ER and PR analysis (FMab vs. RMab) are almost comparable using both detection methods with good overall agreement. For Her2 overexpression, RMab proved to be superior to HerFDA and showed excellent agreement with FISH results with both quantitative detection methods. Diagn. Cytopathol. 2009. © 2009 Wiley‐Liss, Inc.     

Chromogenic in situ hybridization: a novel approach to a practical and sensitive method for the detection of HER2 oncogene in archival human breast carcinoma

The high incidence of HER2 overexpression on the cell surface of breast cancer cells and the recognized prognostic and potentially predictive value of HER2 render this cell surface receptor a novel and important therapeutic target. Although immunohistochemistry (IHC; HercepTest) and fluorescence in situ hybridization (FISH; PathVysion and INFORM)-both approved by the Food and Drug Administration-have emerged as the most viable assays for evaluation of HER2 status in routine clinical practice, there is still no consensus on which is the best method for assessing HER2 status. Therefore, our specific objective was to establish a chromogenic in situ hybridization (CISH) assay for the detection of HER2 amplification on a cohort of 173 archival invasive breast carcinomas. Results were compared with HercepTest, which is the most frequently used method for detecting HER2 alteration. Additionally, HER2 gene copy number was investigated using differential PCR (dPCR) as a testing system. HER2 overexpression was found by IHC in 24.3%; HER2 amplification was found by CISH in 19.1% and by dPCR in 9.2% of the tumors. The overall concordance rate was 95.9% between CISH and IHC and 85.0% between dPCR and IHC. Kappa statistics revealed an excellent agreement between IHC and CISH (kappa = 0.878), but only a moderate agreement was found between IHC and dPCR (kappa = 0.482). Discrepant cases between CISH and HercepTest and all IHC-positive cases (+2 and +3), a total of 42 cases, were analyzed with the FISH PathVysion (Vysis) assay. Among 25 HercepTest-positive cases (score +3), 2 showed no gene amplification by FISH or CISH. Four of 13 tumors with weak HER2 overexpression (score +2) were negative with both FISH and CISH. Concordance between CISH and FISH was 100% for the 38 cases analyzed. The current study showed that CISH represents a practical and simple assay for evaluating HER2 gene amplification in archival material, offering a promising alternative to IHC or FISH for the routine diagnostic setting.     

Role of chromogenic in situ hybridization (CISH) in the evaluation of HER2 status in breast carcinoma: comparison with immunohistochemistry and FISH

We report our experience with Chromogenic in Situ Hybridization (CISH) for the evaluation of HER2 amplification on 55 cases of formalin-fixed, paraffin-embedded invasive breast carcinomas of different histology. All the results were corrected for chromosome 17 aneusomy and compared with immunohistochemistry (IHC); a subset of cases was compared to FISH. Thirty-one of 32 cases in which FISH and CISH were performed yielded the same results. CISH and IHC showed a good concordance in the 0/1+ and 3+ category, while a poor agreement with weakly protein overexpression was confirmed. Chromosome 17 analysis was necessary in cases with a low number of HER2 gene copies. CISH is a useful tool to evaluate breast cancer HER2 status that can be easily implemented in a laboratory of surgical pathology.