Abstract:SUMMARYBACKGROUND. Novel rabbit monoclonal antibodies (RabMab) for estrogen (ER), progesterone (PR) receptors and HER2 evaluation by immunohistochemistry have recently been commercially released. We compared the RabMab anti-ER, anti-PR and anti-HER2 to mouse monoclonal antibodies (Mab) using tissue microarrays (TMA) of breast carcinomas. METHODS. Two TMA containing breast carcinomas were built. Sections were immunostained using anti-ER and anti-PR, Mab and RabMab. The sections stained for ER and PR were evalua… Show more
“…Some series have shown that rabbit monoclonal antibodies (RabMab) for hormone receptors and HER2 have presented better results when compared to mouse monoclonal antibodies (Mabs). However, Mabs for ER (6F11) have shown similar results compared to RabMab and Mabs for HER2 and seemed to be more specific than RabMab for HER2 24 In our study,we evaluated the immunohistochemical staining using an H-score based on intensity and the percentage of stained cells. Semi-quantitative studies for basal markers have established a cutoff of 10% or greater for basal cytokeratin staining to designate the tumor as positive for basal markers 8,9,11 .…”
SummaryObjective. To evaluate the distribution of molecular subtypes of breast tumors diagnosed in young Brazilian women and to analyze the frequency of loss of heterozygocity (LOH) in BRCA1 among different molecular subtypes of early-onset breast cancer. MethOds. Samples from 72 cases of invasive breast carcinoma diagnosed in women aged between 19 and 40 years were evaluated using an immunohistochemical panel of biomarkers. Three intragenic BRCA1 locus microsatellites, D17S1322, D17S1323, and D17S855, were PCR amplified from matched normal (lymphocyte) and tumor DNAs for (LOH) analysis. Results. We found 13 cases (18%) that had an immunohistochemical profile consistent with being basal-like. Forty cases (55%) were luminal A type; 11% (8 cases) were luminal B type, 13% (9 cases) were HER2-overexpressing tumors and two cases were ER-/HER2-carcinomas lacking basal marker expression. Four of the 16 informative cases at D17S1322, one of the four informative cases at D17S855, and none of the five informative cases at D17S1323 displayed LOH (four basal-like and one Luminal A). Microsatellite instability (MSI) at D17S855 and D17S1322 was found in two cases (one a basal-like and one Luminal A). cOnclusiOn. In our study, basal-like tumor was the second most frequent molecular type among young Brazilian women and was only observed in women diagnosed under the age of 35 years. There was no significant difference of LOH at BRCA1 locus rates between basal-like breast tumors and not-basal-like breast tumors (p=0.62). LOH in BRCA1 and MSI in these breast cancers were not frequent but may indicate a small group of breast cancers with a specific molecular makeup.
“…Some series have shown that rabbit monoclonal antibodies (RabMab) for hormone receptors and HER2 have presented better results when compared to mouse monoclonal antibodies (Mabs). However, Mabs for ER (6F11) have shown similar results compared to RabMab and Mabs for HER2 and seemed to be more specific than RabMab for HER2 24 In our study,we evaluated the immunohistochemical staining using an H-score based on intensity and the percentage of stained cells. Semi-quantitative studies for basal markers have established a cutoff of 10% or greater for basal cytokeratin staining to designate the tumor as positive for basal markers 8,9,11 .…”
SummaryObjective. To evaluate the distribution of molecular subtypes of breast tumors diagnosed in young Brazilian women and to analyze the frequency of loss of heterozygocity (LOH) in BRCA1 among different molecular subtypes of early-onset breast cancer. MethOds. Samples from 72 cases of invasive breast carcinoma diagnosed in women aged between 19 and 40 years were evaluated using an immunohistochemical panel of biomarkers. Three intragenic BRCA1 locus microsatellites, D17S1322, D17S1323, and D17S855, were PCR amplified from matched normal (lymphocyte) and tumor DNAs for (LOH) analysis. Results. We found 13 cases (18%) that had an immunohistochemical profile consistent with being basal-like. Forty cases (55%) were luminal A type; 11% (8 cases) were luminal B type, 13% (9 cases) were HER2-overexpressing tumors and two cases were ER-/HER2-carcinomas lacking basal marker expression. Four of the 16 informative cases at D17S1322, one of the four informative cases at D17S855, and none of the five informative cases at D17S1323 displayed LOH (four basal-like and one Luminal A). Microsatellite instability (MSI) at D17S855 and D17S1322 was found in two cases (one a basal-like and one Luminal A). cOnclusiOn. In our study, basal-like tumor was the second most frequent molecular type among young Brazilian women and was only observed in women diagnosed under the age of 35 years. There was no significant difference of LOH at BRCA1 locus rates between basal-like breast tumors and not-basal-like breast tumors (p=0.62). LOH in BRCA1 and MSI in these breast cancers were not frequent but may indicate a small group of breast cancers with a specific molecular makeup.
“…Single FFPE sections of human tissue were stained with 2 to 5 different commercially available, fluorophore-conjugated antibodies against the same protein target (Table 1 ), and the signal intensity from the different clones was compared. The performance of fluorophore-conjugated antibodies was evaluated against the clinical-grade antibodies used in practice by the Pathology Department at Brigham and Women’s Hospital (BWH) 44 . Fig.…”
Emerging data suggests that HER2 intratumoral heterogeneity (ITH) is associated with therapy resistance, highlighting the need for new strategies to assess HER2 ITH. A promising approach is leveraging multiplexed tissue analysis techniques such as cyclic immunofluorescence (CyCIF), which enable visualization and quantification of 10–60 antigens at single-cell resolution from individual tissue sections. In this study, we qualified a breast cancer-specific antibody panel, including HER2, ER, and PR, for multiplexed tissue imaging. We then compared the performance of these antibodies against established clinical standards using pixel-, cell- and tissue-level analyses, utilizing 866 tissue cores (representing 294 patients). To ensure reliability, the CyCIF antibodies were qualified against HER2 immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) data from the same samples. Our findings demonstrate the successful qualification of a breast cancer antibody panel for CyCIF, showing high concordance with established clinical antibodies. Subsequently, we employed the qualified antibodies, along with antibodies for CD45, CD68, PD-L1, p53, Ki67, pRB, and AR, to characterize 567 HER2+ invasive breast cancer samples from 189 patients. Through single-cell analysis, we identified four distinct cell clusters within HER2+ breast cancer exhibiting heterogeneous HER2 expression. Furthermore, these clusters displayed variations in ER, PR, p53, AR, and PD-L1 expression. To quantify the extent of heterogeneity, we calculated heterogeneity scores based on the diversity among these clusters. Our analysis revealed expression patterns that are relevant to breast cancer biology, with correlations to HER2 ITH and potential relevance to clinical outcomes.
The Update Committee recommends that HER2 status (HER2 negative or positive) be determined in all patients with invasive (early stage or recurrence) breast cancer on the basis of one or more HER2 test results (negative, equivocal, or positive). Testing criteria define HER2-positive status when (on observing within an area of tumor that amounts to > 10% of contiguous and homogeneous tumor cells) there is evidence of protein overexpression (IHC) or gene amplification (HER2 copy number or HER2/CEP17 ratio by ISH based on counting at least 20 cells within the area). If results are equivocal (revised criteria), reflex testing should be performed using an alternative assay (IHC or ISH). Repeat testing should be considered if results seem discordant with other histopathologic findings. Laboratories should demonstrate high concordance with a validated HER2 test on a sufficiently large and representative set of specimens. Testing must be performed in a laboratory accredited by CAP or another accrediting entity. The Update Committee urges providers and health systems to cooperate to ensure the highest quality testing. This guideline was developed through a collaboration between the American Society of Clinical Oncology and the College of American Pathologists and has been published jointly by invitation and consent in both Journal of Clinical Oncology and the Archives of Pathology & Laboratory Medicine.
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