HER2 Testing In Breast Cancer Research Articles

Comparative Analysis of Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer According to 2007 and 2013 American Society of Clinical Oncology/College of American Pathologists Guideline Recommendations

Purpose To study the effect of the 2013 updates to the 2007 American Society of Clinical Oncology/College of American Pathologists recommendations for human epidermal growth factor receptor 2 (HER2) testing in breast cancer on testing patterns and interpretation in a large regional reference laboratory. Patients and Methods Patient cases with HER2 testing scores for breast biomarker evaluation were selected from our laboratory information system during two 12-month periods (2012 and 2014). The number of tests performed, type of specimens, proportion of HER2-positive and equivocal patient cases, and number of repeat tests on subsequent excisional specimens were examined and compared. Results Although the number of samples tested increased between 2012 and 2014 (2,201 v 2,558 patient cases; 2,278 v 2,659 tumors), HER2 positivity remained constant (15.7% v 15.5%, respectively). The number of repeat tests performed within 6 months more than doubled (122 [5.5%) of 2,201 v 302 [11.8%] of 2,558; P < .001), and the proportion of immunohistochemistry (IHC) 2+ tumors was significantly lower in 2014 than in 2012 (20.3% v 25.3%; P < .001). However, the proportion of patient cases with unresolved HER2 statuses (equivocal by IHC and in situ hybridization) was significantly higher in 2014 (four of 2,278 v 90 of 2,660; P < .001). Conclusion Our findings indicate that the 2013 updates to the American Society of Clinical Oncology/College of American Pathologists recommendations for HER2 testing in breast cancer did not affect the overall HER2-positivity rate or the proportion of patients eligible for HER2-targeted therapy. The proportion of tests and repeat tests performed increased, as did the number of patient cases categorized as ISH equivocal. The benefit of targeted therapy in the equivocal group is not proven, so targeted therapy should not be considered for patients in this category which should be redefined in future iterations of the recommendations.

Abstract P1-03-08: Simultaneous analyses of HER2 gene and protein status can more precisely predict pathological complete response (pCR) to neoadjuvant trastuzumab with chemotherapy in primary HER2-positive breast cancer

Abstract Background: HER2-positive breast cancer is characterized by overexpression of HER2 protein and/or amplification of the HER2 gene, assessed by immunohistochemistry (IHC) and in situ hybridization (ISH), respectively. According to the 2013 ASCO/CAP guidelines on HER2 testing in breast cancer, both protein and gene status can be classified into three categories: positive, equivocal and negative. Gene-protein assay (GPA) is a newly developed technique, in which IHC and dual color ISH (DISH) are simultaneously performed on a single slide. HER2 GPA slides allow bright-field analyses of both protein and gene expression status of each cell. Breast cancer cells can be classified into nine types based on the combination of protein and gene expression with GPA. Table 1. Three by three classification of cancer cells by the HER2 GPA analysis HER2 protein 3+ (positive)2+ (equivocal)0, 1+ (negative)HER2 gene copy number≥6 (positive)ABC 4, 5 (equivocal)DEF &amp;lt;4 (negative)GHI Purpose: We aimed to elucidate the relationship between the results of HER2 GPA analysis and the therapeutic effects of trastuzumab-based treatment. Methods: Fifty three (53) patients with HER2-positive breast cancer, who underwent neoadjuvant trastuzumab with chemotherapy, were analyzed for HER2 status and clinical outcome. First, HER2 protein and gene status in a pre-therapeutic biopsy material from each patient were separately assessed according to the ASCO/CAP guidelines using HER2 GPA slides. Second, all cancer cells in five representative microscopic images of GPA slides were assessed to determine protein expression and to count the gene copy number at individual cell levels. Finally, we investigated the relationship between the mixture composition of the nine cell types and pathological complete response (pCR) of trastuzumab-treated breast cancer. Results: The GPA results were concordant with the results of IHC in 98% of samples, and with DISH results in 100%. Two hundred eighty nine (289) cancer cells per patient were assessed on average, ranging from 137 to 490 cells. The mean proportion of type A cancer cells was 64%, ranging from 0% to 98%. Patients who had 50% or more of type A cancer cells achieved pCR in 72% of cases (28/39), while patients with less than 50% of type A cancer cells achieved pCR in 7% of cases (1/14) (P&amp;lt;0.0001). The mean proportion of type C cancer cells was 7%, ranging from 0% to 60%. Patients who had 10% or more of type C cancer cells achieved pCR in 10% of cases (1/10), while patients with less than 10% of type C cancer cells achieved pCR in 65% of cases (28/43) (P=0.0016). Conclusion: HER2 GPA analyses can more precisely predict therapeutic effect of trastuzumab-based treatment in primary HER2-positive breast cancer. Our study suggests that the HER2 GPA approach would further improve precision therapy in HER2-positive breast cancer. Citation Format: Horii R, Nitta H, Ito Y, Iwase T, Ohno S, Akiyama F. Simultaneous analyses of HER2 gene and protein status can more precisely predict pathological complete response (pCR) to neoadjuvant trastuzumab with chemotherapy in primary HER2-positive breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-03-08.

Evolution of HER2 testing practices in breast cancer in France from 2005 to 2015: Results of the PartenHER national survey.

e12082Background: For many years, French pathologists have been striving to standardize their practices and improve the quality of HER2 testing in breast cancer. The PartenHER survey aimed to assess how practices have evolved over the last decade. Methods: The PartenHER national survey is based on a questionnaire developed by a scientific committee and sent by postal mail in February 2015. Each French anatomo-cytopathology (ACP) labs received only one questionnaire. The formular was composed of 41 questions including typology of ACP labs, different technical conditions observed during pre-analytical, analytical, post-analytical steps, difficult-to-classify HER2 test results, quality controls, staff organization and training. Results: By 30 July 2015, 109 ACP labs (53% private, 47% public) regularly testing for HER2 and representing almost 620 pathologists (i.e. one third of the French pathologists) had completed the questionnaire. The study showed that in 2015, the immunohistochemistry (IHC) technique was...

The combinatorial approach of laser-captured microdissection and reverse transcription quantitative polymerase chain reaction accurately determines HER2 status in breast cancer.

BackgroundHER2 expression in breast cancer correlates with increased metastatic potential, higher tumor recurrence rates and improved response to targeted therapies. Fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) are two methods commonly used for the analysis of HER2 in the clinic. However, lack of standardization, technical variability in laboratory protocols and subjective interpretation are major problems associated with these testing procedures.MethodsHere we evaluated the applicability of reverse-transcription quantitative polymerase chain reaction (RT-qPCR) for HER2 testing in breast cancer. We tested thirty formaldehyde-fixed and paraffin-embedded tumor samples by RT-qPCR, FISH and IHC and analysed and compared the data from the three methods.ResultsWe found that laser-captured microdissection is essential for the accurate determination of HER2 expression by RT-qPCR. When isolating RNA from total tumor tissue we obtained a significant number of false negative results. However, when using RNA from purified cancer cells the RT-qPCR data were fully consistent with FISH and IHC. In addition we provide evidence that ductal carcinomas might be further classified by the differential expression of HER3 and HER4.ConclusionsLaser-captured microdissection in combination with RT-qPCR is a precise and cost-effective diagnostic approach for HER2 testing in cancer. The PCR assay is simple, accurate and robust and can easily be implemented and standardized in clinical laboratories.

Assessment and significance of human epidermal growth factor receptor 2 gene status in breast cancer with polysomy of chromosome 17

To explore the impact of polysomy of chromosome 17 on the interpretation of human epidermal growth factor receptor 2 (HER2) status and its clinical significance in breast cancers. Clincopathological data of 338 breast cancer patients were collected, and the association of clinicopathological characteristics with polysomy of chromosome 17 was analyzed. Based on the American Society of Clinical Oncology /College of American Pathologists updated recommendations of HER2 testing in breast cancer in 2013, the relationships between the copy number of centromeres on chromosome 17 (CEP17) and HER2 status, other important biological markers and clincopathological features of breast cancer were analyzed statistically. Immunohistochemical examination revealed that among the 338 breast cancer cases, there were 9 (2.7%) HER2-negative cases, 309 (91.4%) HER2-equivocal cases, and 20 (5.9%) HER2-positive cases. The FISH analysis showed that there were 226 (66.9%) HER2-negative cases, 13 (3.8%) HER2-equivocal cases, and 99 (29.3%) HER2-positive cases. Among the 338 breast cancer patients, 42 (12.4%) cases were identified as chromosome 17 polysomy (polysomy 17), 291 (86.1%) cases were chromosome 17 disomy, and 5 (1.3%) cases were chromosome 17 monosomy. Polysomy 17 showed significant correlation with the intensity of HER2 protein expression (P=0.046). The HER2 FISH results and HER2 gene copy number of polysomy 17 cases showed significant differences from the non-polysomy 17 cases (P<0.001). There were no significant differences of HER2/CEP17 ratio between the polysomy 17 and non-polysomy 17 groups (P=0.930). Polysomy 17 cases showed a significant relationship with tumor grading and Ki-67 index (P<0.05), but not with the age of patient, tumor size, lymph node metastasis, ER and PR expressions, and effect evaluation of neoadjuvant chemotherapy (P>0.05). In the breast cancer with polysomy of chromosome 17, assessment of the absolute copy number of HER2 signals is even more important, and the HER2/CEP17 ratio and HER2 protein expression levels should be comprehensively evaluated together.

Comparison of fluorescence in-situ hybridisation with dual-colour in-situ hybridisation for assessment of HER2 gene amplification of breast cancer in Hong Kong.

To compare the PathVysion fluorescence in-situ hybridisation assay with the INFORM HER2 Dual in-situ hybridisation assay on 104 invasive breast cancers with a broad spectrum of immunohistochemistry scores. This case series involved consecutive patients diagnosed with invasive breast carcinoma with equivocal immunohistochemistry score and referred for further HER2 assessment from the departments of Surgery and/or Clinical Oncology of the two hospitals between January 2013 and February 2014. An additional 10 cases with negative HER2 immunohistochemistry and 11 cases with positive HER2 immunohistochemistry were further included. The results of both fluorescence in-situ hybridisation and dual in-situ hybridisation were available in 99 of 104 cases, respectively. Student’st test showed no statistically significant difference in the mean number of HER2 count, CEP17 copies, or HER2/CEP17 ratio between that obtained by fluorescence in-situ hybridisation and that obtained by dual in-situ hybridisation. Pearson’s correlation of results for the two assays was strong for HER2/CEP17 signal ratio (R=0.963, P<0.001) and mean HER2 copies per nucleus (R=0.897, P<0.001). Overall agreement was 96.0% (95 out of 99 cases, ĸ0.882). Three of the four discordant cases were equivocal for either fluorescence in-situ hybridisation or dual in-situ hybridisation. The results of immunohistochemistry 0/1+ and 3+ cases showed 100% concordance between the two assays. The failure rate was 0.96% for fluorescence in-situ hybridisation and 3.85% for dual in-situ hybridisation. Cases that failed for fluorescence in-situ hybridisation were successful for dual in-situ hybridisation and vice versa. Our study showed that dual in-situ hybridisation is a reliable and useful option for HER2 testing in breast cancer.

Application of the 2013 ASCO/CAP guideline and the SISH technique for HER2 testing of breast cancer selects more patients for anti-HER2 treatment.

The aim of this study is to assess the impact of changes of the 2013 ASCO/CAP guideline on the results of HER2 testing in breast cancer. A series of 916 primary invasive breast cancer cases, assessed as HER2 2+ by IHC in part using the 2007 and in part the 2013 ASCO/CAP criteria, was evaluated for HER2 amplification status by SISH and classified according to both 2007 and 2013 ASCO/CAP ISH guideline criteria. We observed a significant increase of HER2-positive cases (12.4 to 16.8%) and a decrease of HER2-equivocal cases (3.6 to 0.7%). Of the cases studied, 52.1% fulfilled both criteria of HER2/CEP17 ratio and average HER2 copy number per cell to be classified as HER2-positive. Reclassification of the cases from before the introduction of the new ASCO/CAP guideline with the 2013 ISH criteria resulted in an increase of cases with a HER2-positive status (12.4 to 14.2%) and in a decrease of HER2-equivocal cases (3.6 to 1.6%). The 2013 ASCO/CAP guideline selects more patients for anti-HER2 targeted therapy, mostly based on the modifications of criteria to evaluate ISH-HER2.

Impact of updated HER2 testing guidelines in breast cancer—re-evaluation of HERA trial fluorescence in situ hybridization data

Recently the American Society of Clinical Oncology and the College of American Pathologists have updated their clinical practice guidelines for HER2 testing in breast cancer. In order to evaluate these new recommendations, we have re-assessed the HER2 status of 6018 breast cancer cases of the screening population for the HERceptin adjuvant (HERA) trial that were originally centrally tested by fluorescence in situ hybridization based on the FDA-released test guidelines. According to the most recent 2013 ASCO/CAP recommendations, 3380 (56.2%) cases were classified as HER2 positive compared with 3359 (55.8%) applying the HERA/FDA scheme and 3339 (55.5%) applying the 2007 ASCO/CAP guidelines. Twenty-one cases switched from negative (HERA/FDA scheme) to positive (2013 ASCO/CAP guidelines). This group is characterized by a mean HER2 gene copy number of ≥6.0, polysomy or co-amplification of CEP17 with an average CEP17 count of 5, and with HER2 receptor overexpression in 75% of cases. On the basis of the HER2 gene copy number alone, we observe 494 cases (8.2%) that are in the equivocal range. Most of these cases (>80%) were also nondecisive by immunohistochemistry (score 2+) irrespective of whether ratio was <2.0>. The number of equivocal cases that would require HER2 reflex testing decreases to 113 (1.9%) if in addition to the HER2 gene copy number also the ratio of HER2 and CEP17 copy numbers is considered via dual-color in situ hybridization. The combination of applying the HER2 mean gene copy number as well as the HER2/CEP17 ratio to define equivocal test decisions by fluorescence in situ hybridization as proposed by the current ASCO/CAP guidelines appears to be a more optimum approach to adopt in order to avoid or minimize reporting of false negative results. Using the mean HER2 gene copy number alone for decision making results in a significant increase of equivocal cases.

New ASCO/CAP guideline recommendations for HER2 testing increase the proportion of reflex in situ hybridization tests and of HER2 positive breast cancers.

Accurate determination of tumour human epidermal growth factor receptor type 2 (HER2) status is critical for optimal treatment of breast cancer. In October 2013, the American Society of Clinical Oncology (ASCO) and the College of American Pathologists (CAP) issued joint updated guideline recommendations for HER2 testing in breast cancer, with a revised algorithm for interpretation of immunohistochemistry (IHC) and in situ hybridisation (ISH) results. This study investigates the impact on HER2 IHC categorisation, implication for reflex ISH testing and potential for identification of false negative IHC. HER2 IHC preparations on 251 invasive breast tumours, originally reported according to 2007 guidelines, were re-scored using 2013 guidelines and the diagnostic categories compared. The results of ISH testing on a separate cohort of 32 breast tumours reported as HER2 IHC 2+ following the introduction of the 2013 guidelines, that would have been designated 1+ according to 2007, were reviewed. Application of 2013 guidelines resulted in a decrease in tumours classified as HER2 negative (83/251 vs 144/251) and a comparable increase in those classified as equivocal (2+) (139/251 vs 80/251). Relatively few tumours were re-classified as positive (29/251 vs 27/251). Furthermore, 3/32 breast cancer cases (HER2 IHC 2+ as per 2013 guidelines, 1+ using 2007 guidelines) were HER2 ISH positive. Application of the 2013 guidelines increases the HER2 IHC equivocal (2+) category and requirement for reflex ISH testing. The reduced threshold for ISH testing identifies some patients with HER2 positive breast cancer whose tumours would have been categorised as HER2 negative according to the 2007 guidelines.

Impact of Modified 2013 ASCO/CAP Guidelines on HER2 Testing in Breast Cancer. One Year Experience.

IntroductionThe latest guidelines of the American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) to test Human Epidermal Growth Factor Receptor 2 (HER2) in breast cancer after being revised in 2008 underwent a second modification in October 2013. The modification includes changes in cut-offs: 10% strong membranous staining for score 3+ on immunohistochemistry (IHC) (previously 30%) and using the ratio of >2 or absolute gene-copy-number (6 or more) alone or in combination with each other by in-situ-hybridisation technology (previously >2.2 and average copy-number of 6 or more). Hereby we addressed the question, which impact the modified cut-offs had on overall HER2-positivity in a single institution.MethodsWe prospectively analysed 617 consecutive diagnostic breast-cancer cases which underwent double HER2 testing by immunohistochemistry and fluorescent in-situ hybridisation (FISH), using the modified 2013 ASCO/CAP-guidelines for one year (October 2013–October 2014). Results were compared with HER2-test results on 1,528 consecutive diagnostic breast-cancer cases from two previous years (2011–2012), using the 2008 ASCO/CAP guidelines, also tested with IHC and FISH in each case.ResultsBetween October 2013 and October 2014, overall HER2-positivity was 15.8% (98 of 617 cases were either IHC 3+ or FISH amplified). 79 of 617 cases (13%) were IHC 3+, 96 of 617 cases (15.5%) were FISH amplified. Equivocal cases were seen in 25 of 617 cases (4.1%). 22 of 25 equivocal cases (88%) in 2013–2014 were IHC 1+ or 2+. In 13 equivocal cases, there was a repeated IHC/FISH testing: 2 of 13 cases (15%) became FISH amplified, 1 of 13 cases (7.5%) became IHC 3+. In 2011–2012, overall HER2-positivity (IHC/FISH) was 13.8% (211 of 1,528 cases). 185 of 1,528 cases (12%) were 3+ on IHC, 181 of 1,522 cases (12%) were amplified by FISH. Six of 1,528 cases were equivocal by FISH, and interpreted as non-amplified (0.3%).ConclusionsApplying the modified ASCO/CAP guidelines from 2013 resulted in an increase (2%) in overall HER2-positivity rate compared to overall-HER2-positivity rate using the 2008 ASCO/CAP guidelines. The increased positivity rate was mainly due to more FISH-positive cases (3.5% more than until 2013). The high rate of equivocal cases (4.1%) did not contribute to increase in overall HER2-positivity, but resulted in delay in definitive HER2-status.

Understanding Current Attitudes to HER2 Testing for Breast Cancer at Tertiary Referral Hospitals of Metro Manila, Philippines

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Current situation and development of HER-2 testing in breast cancer

Current situation and development of HER-2 testing in breast cancer

Assessing HER2 testing quality in breast cancer (BC): Variables that influence HER2-positivity from a large, multicenter, observational study in Germany.

11062 Background: Despite > 10 years of routine HER2 testing in BC, quality is still an issue. It is often assumed that inter- and intra-laboratory variations in testing quality can be assessed by ...

P240 Impact of modified 2013 ASCO/CAP guidelines on HER2 testing in breast cancer

P240 Impact of modified 2013 ASCO/CAP guidelines on HER2 testing in breast cancer

Use of the new 2013 ASCO/CAP guidelines to study HERS status by fluorescence in situ hybridization in patients with breast cancer

Despite the long-term history of HER2 testing in breast cancer (BC), the results of its study are frequently interpreted ambiguously. The introduction of the new 2013 ASCO/CAP guidelines is aimed to further optimize HERS testing in view of new scientific evidence for the biological characteristics of BC. to study the results of assessing HERS2 status in patients with the questionable level of protein expression according to the 2007 versus 2013 ASCO/CAP criteria. Biopsy specimens from 572 patients diagnosed with BC after immunohistochemical examination, including HER2 status assessment, were investigated by fluorescence in situ hybridization. The use of the new 2013 ASCO/CAP criteria versus the previous criteria to determine intratumor heterogeneity, HER2 gene copy number, and the so-called chromosome 17 polysomy was ascertained to cause a statistically significant increase in HER2-positive cases from 25 to 42.5% (OR=2.2178; 95% CI, 1.8254-2.6950; р=0.0005). The use of the new guidelines additionally permits effective therapy in patients with BC, which may be another step to improve survival in this large patient group.

Changing pattern for HER2 positivity due to updated ASCO/CAP guidelines for HER2 testing and its impact.

523 Background: ASCO/CAP guidelines for HER2 testing in breast cancer were published in 2007 and updated in 2013. HER2-directed therapies (HDT) should be recommended in patients who are deemed HER2...

An alternative and sensitive method based on LCM and Q-PCR for HER2 testing in breast cancer.

Nowadays, HER2 testing in breast cancer represents a necessity for both prognostic and therapy. Despite widespread use of immunohistochemistry (IHC) for assessing HER2 status, there are some limitations to identify truly negative or positive HER2 cases. Fluorescence in situ hybridization (FISH) or chromogenic in situ hybridization (CISH) could solve the equivocal HER2 IHC cases but there is no consensus on which is the best method. Consequently, finding a sensitive method for HER2 testing is critical for the management of the disease. In addition, tumor heterogeneity is an important factor which could affect accuracy of molecular diagnostics. Laser capture micro-dissection (LCM) is used to isolate pure cell populations from heterogeneous tumor tissue. The combination between LCM and quantitative polymerase chain reaction (Q-PCR), the gold standard in molecular biology for quantifying gene amplification levels, could define an important tool to improve the molecular diagnostics of HER2 status.In our pilot study we used LCM and Q-PCR to evaluate HER2 gene amplification for invasive breast carcinoma samples. The samples were selected based on HER2 status assessed by IHC and CISH. Our results demonstrated high sensitivity of Q-PCR for assessing HER2 DNA amplification as well as a good concordance between Q-PCR and IHC/ CISH assay.

HER2 Testing in Breast Cancer

Sponsoring Organization: The American Society of Clinical Oncology (ASCO)/College of American Pathologists (CAP)Target Population: Oncologists,

False positivity in HER2 testing of breast cancer: novel paths for approaching an old dilemma

AimsVariability in determining HER2 status has been reported, especially, differences in sensitivity and specificity among commercially available antibodies, with false positive and false negative results. We compared the sensitivity and...

What are NCI-designated cancer centers using for breast cancer HER2 testing?

626 Background: After 15 years of HER2 testing in breast cancer, various testing methods are used in practice. Studies comparing methods are published but no consensus exists on which method is best. Our study provides a benchmark on the use of breast HER2 testing methods in leading U.S. cancer centers. Methods: We conducted an IRB-approved web survey of 58 NCI cancer centers (pathologists and oncologists) providing adult breast cancer care. The survey included 14 questions on breast HER2 testing methods, and reflex and retest practices. We analyzed results using simple frequencies and Fisher's exact test. Results: We achieved a response rate of 98% (57/58 sites). In this cohort, 42% (24/57) of sites conduct HER2 testing for breast cancer using an IHC method with reflex to FISH. Of these, all sites reflex IHC 2+ results and 54% (13/24) automatically reflex IHC results beyond 2+ (see table, results are not mutually exclusive). Concurrent primary FISH and IHC testing is conducted at 32% (18/57) of sites; FISH only testing at 18% (10/57); concurrent primary SISH and IHC testing at 5% (3/57); concurrent primary CISH and IHC testing at 2% (1/57), and CISH only testing at 2% (1/57) of sites. The choice of testing protocol had no correlation with the size of institution, metro vs rural location, NCCN membership, or whether the site acts as a reference lab. However, sites where oncologists always or often request a specific test method were more likely (75%, 21/28) to use FISH as a primary method vs as a reflex method (38%, 9/24), p=.0108. Repeat HER2 testing on surgical tumor samples, after the core biopsy, was reported by 47% of sites (27/57); retesting of relapsed patients by 63% (36/57); and retesting for progressive metastatic disease by 56% (32/57). Conclusions: For HER2 breast biomarker testing, concurrent FISH and IHC testing and expanded reflex testing, beyond IHC 2+ results, have become a common practice at the NCI designated cancer centers. [Table: see text]