Estrogen Receptor Immunohistochemistry Research Articles

The correlation between Ki 67 and Oncotype Dx on HR+, HER2- early breast cancer management: Insights of molecular testing.

e12546 Background: Elevated Ki-67 expression serves as an indicator of tumoral proliferation in breast cancer (BC) and is used in Ecuador for survival estimation, recurrence risk assessment, and guiding chemotherapy decisions. The Oncotype DX Breast Recurrence Score (ODX RS) is a validated 21-gene assay for hormone receptor-positive, HER2-negative invasive BC. It predicts chemotherapy benefits and the risk of distant recurrence over 10 years and is a mortality indicator. ODX RS has the potential to reduce the need for chemotherapy, alleviate financial burdens, and enhance clinical outcomes. Despite conflicting evidence on the significance of Ki-67 in the ODX RS, there is a lack of exploration within Latin populations. This study aims to compare the correlation of immunohistochemical (IHC) measured Ki-67 to the ODX RS in the Ecuadorian population. Methods: A retrospective analysis included women with early BC who underwent ODX RS testing in Ecuador from May 2020 to January 2023. Inclusion criteria: early BC (T1-T3/N0-N1a); IHC estrogen receptor (ER) positive; HER2 negative; and availability of tumor size, grade, lymph node status, and Ki67 proliferation index data. Exclusion criteria: patients with recurrent BC, incomplete pathology, or lacking IHC data. Patients were categorized into low (≤5%), intermediate (6%–29%), and high Ki67 expression groups (≥30%). Using the statistical software SPSS, the correlation between ODX scores and Ki-67 expression was analyzed using Pearson’s correlation coefficient. For risk stratification groups, Spearman's correlation was employed. Results: Among 77 patients undergoing Oncotype DX testing, 64 (with a mean age of 55) met the inclusion criteria. Eighty-six percent of cases were invasive ductal carcinomas (IDC). Seventeen percent exhibited low Ki-67 expression, 61% intermediate, and 20% high. No statistically significant correlation was found between IHC Ki-67 expression and ODX RS (r=0.051; p=0.69), neither when stratified by categories (r=0.086; p=0.50). A subanalysis based on risk categories showed 82% correspondence in the low Ki-67 group, 18% in the intermediate group, and 31% in the high Ki-67 group. No significant correlation was observed when stratified by histological types (invasive ductal carcinoma and invasive lobular carcinoma), nor by sample origin (core biopsy, excisional biopsy). Conclusions: There was no correlation between IHC Ki-67 expression and ODX risk scores in this study involving Ecuadorian patients. Our findings emphasize that IHC Ki-67 expression should not be the sole determinant for guiding therapeutic decisions and predicting recurrence in early breast cancer.[Table: see text]

Novel epigenomic liquid biopsy assay to predict estrogen receptor (ER) status and to infer ER pathway activation in breast cancer.

1066 Background: Endocrine therapy (ET) is the cornerstone of treatment for ER-positive (ER+) breast cancer (BC). However, BC may lose ER expression over the disease course, and ER+ tumors may become resistant to ET regardless of ER expression. Blood-based biomarkers to infer ER expression and to predict ET sensitivity are lacking. Methods: We identified 44 patients (pts) with metastatic BC seen at Dana-Farber Cancer Institute from 2012-2023, who had blood samples drawn in Streck tubes within 6 weeks of tumor biopsy with ER scored via immunohistochemistry (IHC). We applied a novel, multi-analyte, liquid biopsy assay to capture genome-wide epigenomic signals, mapping enhancers, promoters, and DNA methylation data from 1mL of plasma. First, we applied a previously developed, IHC-benchmarked, regularized logistic regression model to infer ER status from plasma-based epigenomic profiles (Parsons et al., SABCS 2023). Second, we applied a novel ER pathway activity score involving promoters and enhancers of certain genes previously associated with ER activity (Guan et al., Cell 2019). The ER activity score was corrected for the circulating tumor DNA (ctDNA) fraction as assessed by low pass whole genome sequencing (ichorCNA algorithm). We evaluated the association between the ER activity score and the ER status by IHC using the Wilcoxon rank-sum test. ER expression ≥1% was considered positive. Results: The ER classifier and the ER activity score have been applied to 19 of 44 samples (43%) all of which had detectable ctDNA. Eleven out of 19 (58%) pts had ER+ BC by IHC (ER IHC range 80-95%). Overall, 18/19 (95%) were correctly classified as ER+ or ER- by the ER classifier (AUC=1). Three of 19 (16%) samples were collected from pts with ER+ BC at primary diagnosis and ER- BC at time of tissue sampling. For them, the ER classifier correctly predicted ER- status. We observed a significantly higher ER activity score in samples from pts with ER+ BC than pts with ER- BC as measured by IHC (p= 0.0012). For pts with ER+ BC, we observed an association between lower ER activity scores and a higher number of lines of ET prior to blood sampling, with lower scores for pts who received >4 prior lines. We will present full results from this cohort and an expanded cohort and correlate the ER score with response to ET. Conclusions: We applied a novel blood-based epigenomic classifier able to infer ER status and a novel epigenomic activity score to predict ER signaling from a minimal plasma volume. We plan to validate our findings on a larger cohort and correlate with response to ET. If confirmed, this assay may represent a valuable, minimally invasive tool to help guide treatment selection for patients with ER+ BC and predict response to novel endocrine agents.

The role of androgen receptors in breast cancer

BackgroundBreast cancer is a heterogenous group of diseases with steroid hormone dependant carcinogenesis. Along with Estrogen Receptor (ER), Progesterone Receptor (PR), Human Epidermal growth factor Receptor 2 (HER2), androgen receptor (AR) is now emerging to play key role in disease progression of various molecular breast cancer subtypes especially triple negative breast cancers. Aim of this study was to find out prevalence of AR positivity in all breast cancers, correlation of ER, PR, HER2 and Ki67 index with AR positivity. To find out correlation of AR positivity with histological grade. MethodsThis was a descriptive cross-sectional study done on 55 primary breast carcinoma cases. Routine Hematoxylin and Eosin stain as well as immunohistochemical ER, PR, HER2/neu, Ki67 and AR stain were done. The outcome was assessed in the form of relationship between AR and ER, PR, HER2 status and Ki-67 index and histopathological grade. ResultsAR positivity was seen in 69% of all breast cancers. AR positivity was seen in 8% of triple negative breast cancers. High Ki67 index showed 55% AR positivity. Grade II breast cancers showed 42% AR positivity. ConclusionCases with triple negative breast cancers have worst prognosis. Unlike targeted novel hormone therapy (tamoxifen) in luminal cancers and monoclonal antibody (trastuzumab) in her2 enriched cancers, AR is looked upon as a target to treat triple negative breast cancer. However, the existing controversies demand more AR related studies.

Augmented interpretation of HER2, ER, and PR in breast cancer by artificial intelligence analyzer: enhancing interobserver agreement through a reader study of 201 cases

BackgroundAccurate classification of breast cancer molecular subtypes is crucial in determining treatment strategies and predicting clinical outcomes. This classification largely depends on the assessment of human epidermal growth factor receptor 2 (HER2), estrogen receptor (ER), and progesterone receptor (PR) status. However, variability in interpretation among pathologists pose challenges to the accuracy of this classification. This study evaluates the role of artificial intelligence (AI) in enhancing the consistency of these evaluations.MethodsAI-powered HER2 and ER/PR analyzers, consisting of cell and tissue models, were developed using 1,259 HER2, 744 ER, and 466 PR-stained immunohistochemistry (IHC) whole-slide images of breast cancer. External validation cohort comprising HER2, ER, and PR IHCs of 201 breast cancer cases were analyzed with these AI-powered analyzers. Three board-certified pathologists independently assessed these cases without AI annotation. Then, cases with differing interpretations between pathologists and the AI analyzer were revisited with AI assistance, focusing on evaluating the influence of AI assistance on the concordance among pathologists during the revised evaluation compared to the initial assessment.ResultsReevaluation was required in 61 (30.3%), 42 (20.9%), and 80 (39.8%) of HER2, in 15 (7.5%), 17 (8.5%), and 11 (5.5%) of ER, and in 26 (12.9%), 24 (11.9%), and 28 (13.9%) of PR evaluations by the pathologists, respectively. Compared to initial interpretations, the assistance of AI led to a notable increase in the agreement among three pathologists on the status of HER2 (from 49.3 to 74.1%, p < 0.001), ER (from 93.0 to 96.5%, p = 0.096), and PR (from 84.6 to 91.5%, p = 0.006). This improvement was especially evident in cases of HER2 2+ and 1+, where the concordance significantly increased from 46.2 to 68.4% and from 26.5 to 70.7%, respectively. Consequently, a refinement in the classification of breast cancer molecular subtypes (from 58.2 to 78.6%, p < 0.001) was achieved with AI assistance.ConclusionsThis study underscores the significant role of AI analyzers in improving pathologists' concordance in the classification of breast cancer molecular subtypes.

Detection of KRAS Mutations in Triple-negative Breast Cancers by Polymerase Chain Reaction

Abstract Background: Recently, it has been proven that KRAS gene mutation analysis is an additional in vitro diagnostic method for determining colorectal cancer patients who will not respond to anti-epidermal growth factor receptor (EGFR) therapy. KRAS mutation status assessment may be important in other EGFR-overexpressed cancers, such as breast cancer tumors. Only 5% of breast tumors are KRAS-mutated, yet 60% of triple-negative breast tumors express EGFR, making them potential targets for EGFR inhibitors. To provide a biological foundation for assessing anti-EGFR therapy, we aimed to examine the frequency of KRAS mutations in triple-negative breast cancer (TNBC). Materials and Methods: Forty breast cancer cases were evaluated. These 40 cases have undergone immunohistochemistry for estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 markers which exhibited a negative reaction and were ascertained as triple negative. The mutation status of KRAS was determined by real-time polymerase chain reaction (RT-PCR) to detect wild or mutant types of KRAS. Results: In our study, we observed that, out of the total 40 cases of TNBCs, 14 (35%) cases were of wild type and 26 (65%) cases were mutant type detected by RT-PCR. Conclusion: According to our study, KRAS mutations were detected in TNBCs, and EGFR inhibitors may be effective in the treatment of these tumors, which overexpress EGFR in around 65% of cases.

Masculinizing hormone therapy effect on breast tissue: Changes in estrogen and androgen receptors in transgender female-to-male mastectomies

PurposeAlmost two percent of individuals in the United States identify as gender non-conforming. In the female-to-male (FTM) transgender population, masculinizing hormone therapy with testosterone is commonly prescribed in gender transition. To date, the effects of exogenous androgens on breast tissue and its roles in altering breast cancer risk are poorly understood. This study examines the histopathologic findings in gender affirming mastectomy (GAM) in transgender FTM patients and the effects of exogenous androgens on estrogen receptors (ER) and androgen receptors (AR). MethodsA retrospective review of pathology specimens obtained between 2017 and 2020 was performed comparing androgen exposed breast tissue with breast tissue without androgen exposure. Breast specimens were obtained from patients who underwent FTM GAM with recorded exogenous androgen exposure. Control breast specimens were obtained from reduction mammoplasty (RM) procedures in cisgender women which were aged matched to the GAM cohort, as well as postmenopausal women with benign/prophylactic mastectomy procedures; all controls were without androgen exposure. The histopathologic findings were assessed. Immunohistochemistry for AR and ER was performed and the score interpreted by digital image analysis. ResultsAndrogen-exposed breast tissue revealed dense fibrotic stroma, lobular atrophy, thickened lobular basement membranes, and gynecomastoid changes. Longer duration of androgen exposure resulted in a more pronounced effect. The incidence of atypia or cancer was lower in GAM than RM cohort. ER and AR expression was highest in transgender male breast tissue with intermediate duration of exogenous androgen exposure. ConclusionIncreased androgen exposure is associated with lobular atrophy and gynecomastoid changes in breast parenchyma. Overall, ER and AR are expressed strongly in lobular epithelium in patients with prolonged androgen exposure. Exogenous testosterone does not appear to increase risk for breast cancer. Additional studies are needed to investigate the mechanism responsible for these changes at a cellular level and its role in cancer development.

Tonsil tissue control is ideal for monitoring estrogen receptor immunohistochemical staining.

Estrogen receptor (ER) testing performed using immunohistochemistry (IHC) is a critical predictive tool for breast cancer treatment. This study aimed to investigate the use of tonsil control for monitoring ER staining and hypothesize that optimal staining would reduce interlaboratory variations. A proficiency test for ER IHC was conducted using 21 tissue cores. The staining quality was centrally reviewed based on tonsil ER staining. We found that 64.9% of participant samples demonstrated optimal or good staining quality. Poor staining quality was significantly associated with the use of Ventana autostainers and concentrated antibodies. Although the concordance rate did not show significant differences across staining quality levels, interparticipant agreement declined as staining quality deteriorated. Among the 19 discordant responses, 63.2% could be attributed to staining problems, whereas 36.8% could be due to misinterpretation. Poor staining quality due to inadequate staining was the primary reason for undercalls, which can lead to false-negative results. Misinterpretations of nonspecific faint staining that was weaker than the staining of the tonsil control were the cause of most overcalls. Tonsil tissue is an ideal control for monitoring ER staining and can serve as a reference for determining the lower bound for ER positivity. Optimal ER staining and appropriate references for ER positivity can further improve ER IHC quality.

To study the expression of estrogen, progesterone receptor and p53 immunohistochemistry markers in subtyping endometrial carcinoma.

Endometrial cancer is one of the most commonly diagnosed cancers in women worldwide. To study the expression of estrogen receptor (ER), progesterone receptor (PR) and p53 immunohistochemistry (IHC) markers in subtyping endometrial carcinoma. Materials and Methods: A total of 100 cases of carcinoma endometrium submitted during January 2016 to October 2018 were included in our study. The ER, PR and p53 expressions were scored as per the adopted scoring system. Agreement between ER, PR and p53 IHC expression and the consensus HE diagnosis, FIGO grading and tumour staging were assessed using Chi square tests. There was a statistical association between ER, PR and p53 status and tumour histologic type with a P value < 0.01. There was no statistical significance observed between ER and PR expressions and different FIGO grades. Statistical significance (P = 0.036) between p53 and different FIGO grades seen. No statistical significance was observed between ER, PR and p53 expressions and different tumour stages and tumour invasiveness. There was a statistical association between ER and PR status and lymph node metastasis. p53 did not show a statistical significance. Combination of ER, PR and p53 IHC markers can be used to distinguish type 1 and type 2 endometrial cancers. PR expression is more specific than ER in endometrioid carcinomas. p53 expression is more specific in serous carcinoma, however, p53 IHC alone cannot be used to distinguish different grades of endometrioid carcinomas as there is variability of staining in endometrioid carcinomas.

Correlation between Magee score and recurrence score in Indian patients with breast cancer.

e12555 Background: Recurrence score (RS) from Oncotype Dx (ODx) test predict chemotherapy benefit in estrogen receptor (ER) positive, and HER2-negative patients with breast cancer. ODx is an expensive test in an Indian practice setting - cost of adjuvant chemotherapy is one-sixth of the cost of the test. Magee equations use histopathological variables, and semiquantitative results from ER, PR and Ki-67, to calculate Magee score (MS) and provide a cost-effective surrogate mechanism for predicting RS. We aimed to assess the correlation between MS and RS in an Indian setting. Methods: Pathology reports were obtained from oncologists in India. Immunohistochemistry for ER and PR were reported using the Allred system. Therefore, two methods were identified to estimate H-scores - using a calculated method (A), and using an expert-approximated (B) method. MS were calculated by a single investigator blinded to the RS. MS and RS were then assessed for correlation via the Pearson correlation method. MS were arbitrarily stratified into ≤ 15, 15 - 30, and ≥ 30, and their overall agreement (OA) or concordance with RS risk categories, ≤ 15, 15 - 30, and ≥ 30, were assessed. The OA was assessed by determining the proportion of patients in similar risk categories across low, intermediate and high-risk RS and MS. Results: Among 34 patients included in the study, 38% (13) had an RS ≤ 15, 38% (13) had an RS between 15 and 30, and 23% (8) had an RS ≥ 30. Correlation coefficient between MS and RS was 0.78 for both methods of estimation (p&lt;0.0001). The OA between MS-A and RS, and MS-B and RS were 59% (20/34) and 47% (16/34) respectively. The concordance between low-risk MS-A and RS is 78% (n=9) and between low-risk MS-B and RS is 50% (n=20). The concordance between high-risk MS-A and RS is 100% (n=2) and high-risk MS-B and RS is 100% (n=2). Conclusions: Although there is good correlation between RS and MS, the concordance between the two in similar risk categories is not optimal. Of the two methods to estimate H-score from Allred score, the calculated method seems to be more optimal. Our pilot study shows good correlation between MS and RS. We recommend that pathologists incorporate H-score as part of the reporting of ER and PR. A larger prospective study will be useful to test our hypothesis for the use of MS as a predictive tool for adjuvant chemotherapy in breast cancer.

Functional estrogen receptor signal transduction pathway activity and antihormonal therapy response in low-grade ovarian carcinoma.

Advanced low-grade ovarian carcinoma (LGOC) is difficult to treat. In several studies, high estrogen receptor (ER) protein expression was observed in patients with LGOC, which suggests that antihormonal therapy (AHT) is a treatment option. However, only a subgroup of patients respond to AHT, and this response cannot be adequately predicted by currently used immunohistochemistry (IHC). A possible explanation is that IHC only takes the ligand, but not the activity, of the whole signal transduction pathway (STP) into account. Therefore, in this study, the authors assessed whether functional STP activity can be an alternative tool to predict response to AHT in LGOC. Tumor tissue samples were obtained from patients with primary or recurrent LGOC who subsequently received AHT. Histoscores of ER and progesterone receptor (PR) were determined. In addition, STP activity of the ER STP and of six other STPs known to play a role in ovarian cancer was assessed and compared with the STP activity of healthy postmenopausal fallopian tube epithelium. Patients who had normal ER STP activity had a progression-free survival (PFS) of 16.1months. This was significantly shorter in patients who had low and very high ER STP activity, with a median PFS of 6.0 and 2.1months, respectively (p<.001). Unlike ER histoscores, PR histoscores were strongly correlated to the ER STP activity and thus to PFS. Aberrant low and very high functional ER STP activity and low PR histoscores in patients with LGOC indicate decreased response to AHT. ER IHC is not representative of functional ER STP activity and is not related to PFS.

Beyond the estrogen receptor: In search of predictive biomarkers for low-grade serous ovarian cancer.

Estrogen receptor immunohistochemistry should not be used in the clinic as an indicator of response to antihormonal therapy; progesterone receptor immunohistochemistry may be prognostic, but further study is needed. Multigene assays to assess for estrogen receptor pathway activation hold potential as a viable diagnostic tool to help determine those patients least likely to respond to single‐agent endocrine therapy and potentially identify patients most appropriate for combination strategies.

Abstract P3-05-08: Prevalence and prognosis of ER-loss in advanced invasive lobular carcinoma

Abstract Introduction: Estrogen receptor (ER) loss occurs in about 20% of recurrent breast cancers (BC) and is associated with unresponsiveness to endocrine therapy (ET) and poor prognosis. Prior studies evaluating ER-loss included predominately patients with invasive ductal carcinoma (IDC), and therefore the impact of ER-loss in invasive lobular carcinoma (ILC) is unknown. In this retrospective analysis, using real-world data, we aimed to determine the prevalence and clinical significance of ER-loss in ILC. Methods: Advanced BC were molecularly profiled at Caris Life Sciences (Phoenix, AZ) with NextGen Sequencing of DNA (592-gene panel or whole-exome sequencing), RNA (whole transcriptome sequencing, WTS) and immunohistochemistry (IHC) of select markers. A large real-world evidence (RWE) database combining Caris’ molecular data with clinical information obtained from insurance claims data (CODEai) was interrogated and overall survival (OS) was calculated from time of tissue collection to last patient contact. A tumor was considered to have ER-loss if therapies approved only for ER-positive BC were prescribed prior to obtaining a negative ER IHC result. OS was compared using Kaplan-Meier estimates for defined patient cohorts; significance was determined as p values &amp;lt; 0.05. For molecular analyses, Fisher-Exact or Chi-Square tests were used to determine p values. Correction for multiple comparisons was performed using Benjamini-Hochberg to calculate q values. Results: The RWE database included 24,824 patients with advanced BC. At the time of tissue collection for molecular profiling, 6,786 advanced BC patients had been previously treated with ET (with or without mTOR or CDK4/6 inhibitors), of whom 1,338 had data available on histologic classification and ER IHC. The final analytical cohort included 263 patients with ILC and 1,075 with IDC. ER-loss was identified in 11.4% of ILC (n=30/263) and 19.6% (n=210/1075) of IDC (p=0.0017). In ILC, ER-loss was associated with significantly worse OS (HR: 1.75, 95%CI: 1.10-2.79, p=0.016) compared with no ER-loss. In the cohort of patients with ER-loss, patients with ILC had significantly worse OS compared with IDC (HR=2.03, 95% CI: 1.267-3.251, p=0.003). Further, when 1,016 tumors with ER-loss (regardless of histology) were stratified by the median OS (mOS=11mo), positive PD-L1 expression (34% vs. 22%, p=0.04, q=0.22), HER2 IHC positivity (16% vs. 7.8%, p=0.003, q=0.08) and HER2 amplification (16% vs. 4.7%, p=0.0006, q=0.04) were enriched in patients with longer mOS; while amplification of TEFB (0.38% vs. 2.6%, p=0.047, q=0.23) and MYB (0.38% vs. 2.6%, p=0.047, q=0.23) were enriched in patients with shorter mOS. WTS identified 197 differentially expressed genes, the majority of which were enriched in patients with longer mOS (q&amp;lt; 0.05). Conclusions: In this large real-word dataset, ER-loss likely occurred in 11.4% of ILC and was associated with worse OS compared to both patients with IDC and ER-loss and ILC without ER-loss. Our analysis had several limitations; notably, our definition of ER-loss was based on prior treatment, we could not distinguish between de novo or recurrent metastatic disease and time of tissue collection was not standardized during the course of treatment. Thus, additional studies are needed to confirm these findings. However, this study does suggest that ER-loss occurs in a subset of patients with ILC and has poor prognostic implications. Citation Format: Whitney L. Hensing, Joanne Xiu, W. Michael Korn, Stephanie L. Graff, Irene Kang, Evanthia T. Roussos Torres, Arielle L. Heeke, Andrew A. Davis, Nusayba A. Bagegni, Katherine K. Clifton, Ron Bose, Cynthia Ma, Foluso O. Ademuyiwa. Prevalence and prognosis of ER-loss in advanced invasive lobular carcinoma [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-05-08.

Abstract P4-09-10: Spatial protein and RNA expression in tumor, immune and stromal cells in BRCA1/2 mutated metastatic breast cancer

Abstract Background: Spatial transcriptomics and proteomics are cutting-edge techniques that allow for quantification of RNA and protein expression within separate cell populations or regions of interest in a tissue biopsy specimen. We aimed to independently characterize the tumor cells and immune infiltrate in metastatic site biopsy specimens from patients with somatic or germline BRCA1/2 mutations. Methods: We performed spatial transcriptomics and proteomics using the Nanostring GeoMx Digital Spatial Profiler (DSP) on 9 formalin-fixed paraffin-embedded (FFPE) tissue biopsy specimens from 5 patients with metastatic breast cancer and a known somatic or germline BRCA1/2 mutation. Each patient had 1-3 separate metastatic site tissue biopsy specimens. Cell populations of interest were delineated with fluorescently labeled antibodies to pan-cytokeratin on tumor cells, CD45 of immune cells, and a nuclear stain. On each sample, 4-5 regions of interest (ROIs) were selected near the interface of the metastatic tumor deposit and surrounding normal tissue. Each ROI was segmented into 3 areas of interest (AOIs): pan-cytokeratin-positive tumor cells, CD45-positive immune cells, and the remaining stromal cells negative for both pan-cytokeratin and CD45. Data was background corrected and normalized with internal negative and positive internal controls. Relative abundance of immune cell subtypes was derived from CD45+ cell gene expression data using the TIMER algorithm. Results: The expression of 57 proteins and 84 RNAs was quantified in each of the 396 AOIs. Separate analysis of CD45+ cells, pan-cytokeratin cells, and stromal cell populations confirmed expected differential expression of immune, tumor and stromal markers, respectively. Protein expression of the estrogen receptor (ER) and progesterone receptor (PR) on pan-cytokeratin-positive tumor cells using the DSP correlated with pathologist reviewed immunohistochemistry (IHC) for ER and PR. Significant differential expression was found within each cell population between hormone receptor (HR) positive and negative cohorts, including enrichment for S100B in HR negative samples. Comparison will be shown for expression of 14 overlapping genes/proteins. Immune cell compartment gene expression deconvolution showed heterogeneity in the relative proportion of infiltrating immune cell subtypes between samples and also between ROIs within a sample. Changes in gene and protein expression from a patient with 3 serial biopsy specimens can be tracked with substantial heterogeneity across distinct AOIs within individual specimens and across metastatic sites.. Conclusions: Spatial transcriptomics and proteomics is feasible for concurrent, relative quantification of protein and gene expression in tumor, immune, and stromal cells in FFPE tissue biopsies from patients with metastatic breast cancer harboring germline or somatic BRCA1 or BRCA2 mutations. Heterogeneity within individual samples and across serial time points offers opportunity for methods development in the analyses and interpretation of spatial transcriptomic/proteomic data. Citation Format: Katharine A. Collier, Katherine Miller, Zaibo Li, Jesse Westfall, David Tallman, Mark Vater, Gabriel Tinoco, Elaine Mardis, Daniel Stover. Spatial protein and RNA expression in tumor, immune and stromal cells in BRCA1/2 mutated metastatic breast cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P4-09-10.

NOHA: A Promising Biomarker for Determining Estrogen Receptor Status Among Patients With Breast Cancer in Resource-Constrained Settings.

Challenges to breast cancer control in low-and middle-income countries exist because of constrained access to care, including pathology services. Immunohistochemistry (IHC)-based estrogen receptor (ER) analysis is limited-nonexistent because of few and inadequately staffed and equipped pathology laboratories. We have identified Nw-hydroxy-L-Arginine (NOHA) as a blood-based biomarker to distinguish ER status in US patients with breast cancer. Here, we examine NOHA's clinical utility as an ER IHC alternative in Tanzanian patients. Following informed consent, 70 newly diagnosed, known or suspected patients with breast cancer were enrolled at Kilimanjaro Christian Medical Center; basic, deidentified clinical and sociodemographic data were collected. For each, a needle prick amount of blood was collected on a Noviplex plasma card and stored at -80°C. Plasma cards and unstained tumor pathology slides were shipped regularly to US laboratories for NOHA, histologic and IHC analysis. NOHA and IHC assay operators were blinded to each other's result and patient clinical status. Paired NOHA and IHC results were compared. Slides from 43 participants were available for pathological analysis in the United States. Of those with confirmed malignancy (n = 39), 44%, 51%, 5% were ER-positive, ER-negative, and ER inconclusive, respectively. NOHA levels were available among 33 of 43 of those with pathological data and showed distinct threshold levels correlating 100% to tumor ER IHC and disease categorization where a level below 4 nM, from 4 to 8 nM, and above 8 nM signified ER-negative, ER-positive, and no cancer, respectively. The results are consistent with findings from US patients and suggest NOHA's clinical utility as an accessible IHC replacement in determining ER status among low-and middle-income country patients with breast cancer, promising to extend access to cost-efficient, available hormonal agents and improve outcomes.

Role of Estrogen Receptor Alpha rs3798577 Polymorphism in Breast Carcinoma Risk Determination

BACKGROUND: Interaction between estrogen and estrogen receptor (ER) takes part in the regulation and differentiation of breast tumorigenesis. Some ERα polymorphisms, including ERα rs3798577, are reported to be associated with the risk and aggressiveness of breast carcinoma since the site was reported to be targeted by microRNA, which can further modulate the ERα expression. Hence, this study was conducted to disclose the possible role of ERα SNP rs3798577 on breast carcinoma patients.METHODS: Samples were taken from the post-mastectomy breast carcinoma tissues of female patients and screened based on the completeness of medical and histopathological records. DNA isolation was proceeded using real time-polymerase chain reaction (RT-PCR) then analyzed for high resolution melting (HRM). The nucleotide base sequence was then analyzed based on rs3798577 ERα polymorphism. ER immunohistochemistry test was carried out and counted quantitatively based on the staining intensity and the percentage of the stained cells.RESULTS: Out of 65 samples, there were 33 samples as wild type and 32 samples as variant type. Most variant and wild type had &gt;80% ERα percentage. Most variant type had middle ERα intensity, while wild type had strong ERα intensity. Higher percentage of variant type (52.2%) was found with weak ERα histoscore, meanwhile higher percentage of wild type (52.4%) was found with strong ERα histoscore, but not significant (p=0.725).CONCLUSION: ERα rs3798577 variant type had a lower ERα intensity and weaker ERα histoscore compared to the wild type, suggesting that ERα rs3798577 polymorphism might play a role in breast carcinoma risk determination.KEYWORDS: breast cancer, ERα, rs3798577, polymorphism, immunoexpression

Comparison of HER-2/neu with histological grade and hormone (ER, PR) status in carcinoma breast

Objectives :- The study was performed with the aim to compare HER-2/neu over expression with age, size of tumor, histopathological type, grade of tumor, lymph node positivity, NPI, and hormone receptors ER and PR. Methods :- The study was conducted on 66 cases of breast carcinoma. All the cases underwent immunohistochemistry for ER, PR and HER-2/neu over expression. HER-2/neu over expression was compared with prognostic factors. Results :- The age range of these patients was from 30-70 years with mean age of 49.92 years. HER-2/neu was strongly positive (score 3) in 12/66 ( 18.18%), moderate or equivocal ( score 2) in 7 /66 (10.60 %) and mild ( score 1) in 16/66 (24.24%) and it was negative ( score 0) in 31/66 (46.97 %). Estrogen Receptor ( ER) was positive in 38/66 (57.57%) cases, progesterone receptors (PR) was positive in 30/66 (45.45%) cases. Triple negative cases were 16/66 (24.24%) and triple receptor positive cases were 04/66 (6.06%).

Neurologic, functional, and survival outcomes following surgical management of metastatic breast cancer to the spine

ObjectiveMetastatic spinal tumors commonly arise from primary breast cancer. We assessed outcomes and identified associated variables for patients who underwent surgical management for spinal metastases of breast cancer. MethodsWe retrospectively reviewed patients surgically treated for spinal metastases of breast cancer. Neurologic and functional outcomes were analyzed via Frankel scale and Karnofksy Performance Status (KPS) scores, respectively. Variables associated with Frankel and KPS scores after surgery were identified. Multivariable analysis was used to assess predictors for postoperative survival. ResultsForty-nine patients were identified. There was no significant difference in Frankel scores postoperatively and at last follow-up. KPS scores (P = 0.002) significantly improved at last follow-up. Preoperative non-ambulation and postprocedural complications were associated with non-ambulation postoperatively. Postprocedural complications and disease-free interval (DFI) < 24 and < 60 months were associated with functional impairment at last follow-up. Current smoking status at the time of surgery (P = 0.021) and triple negative (negative immunohistochemistry for estrogen receptor, progesterone receptor, and HER2) breast cancer (P = 0.038) were significantly associated with shortened postoperative survival. ConclusionWhen indicated, surgery for spinal metastases of breast cancer leads to preservation of neurologic status and long-term functional improvement. Preoperative ambulatory status and postprocedural complications were associated with ambulatory status after surgery, while postprocedural complications and shortened DFI were associated with functional status after surgery.Current smoking status at the time of surgery and triple negative breast cancer are negative predictors for postoperative survival after metastatic breast cancer to the spine.

To Evaluate the Expression of p53 and Ki67 in Breast Cancer and Their Clinicopathological Correlation with ER PR and HER2/neu

Background: Breast cancer is the most common cancer among women in India and with an increasing trend. The combined estrogen receptor (ER), progesterone receptor (PR), and HER2/neu biomarker expression is a predictor of breast cancer status for therapeutic guidance. Studies have demonstrated that these biomarkers are unstable throughout their tumor progression. Varying concordance and discordance in the biomarker expression between ER, PR Her2neu and its correlation with p53 and ki67 status are reported. Aim: This study was conducted for studying and comparing the expression of immunohistochemistry (IHC) markers, i.e., ER, PR, HER2/neu, p53, and Ki67 for clinicopathological features and interrelation purpose. Methods: Fifty one cases of female breast carcinoma between years December 2019 and April 2021 were included in the study. IHC was applied on the sections from tru-cut/excisional biopsies and subsequent mastectomy specimens. Analyses of the expression of IHC markers were attempted. Results: Substantial agreement was observed for biomarker ER, PR, HER2/neu, p53, and Ki67 expression with respect to age of the patient, size of the lump, histologic grade and diagnosis. Conclusion: This study demonstrated a significant association between tumor size with advanced histological grade, high cell proliferation (Ki-67 expression) and p53. Thus, there is a need to perform ER, PR, HER2/neu, p53, and Ki67 IHC studies routinely to help design therapies that are tailored to target the specific tumor clones and render maximum benefit to patients.

Concordance of breast cancer biomarker testing in core needle biopsy and surgical specimens: A single institution experience.

Accurate diagnostic biomarker testing is crucial to treatment decisions in breast cancer. Biomarker testing is performed on core needle biopsies (CNB) and is often repeated in the surgical specimen (SS) after resection. As differences between CNB and SS testing may alter treatment decisions, we evaluated concordance between CNB and SS as well as associated changes in treatment and clinical outcomes. We performed a retrospective analysis of breast cancer patients at our institution between January 2010 and May 2020. Concordance between CNB and SS was assessed for estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). Survival in patients, including recurrence, metastatic recurrence, and death, were assessed using chi-squared likelihood ratio. In total, 961 patients met eligibility criteria. Concordance, minor discordance, total concordance (concordance plus minor discordance), and major discordance between CNB and SS were reported for ER (87.7%, 9.2%, 90.8%, and 2.9%), PR (58.1%, 29.1%, 87.2%, and 12.8%), and HER2 IHC (52.5%, 20.9%, 73.4%, 26.6%), respectively. HER2 FISH concordance and major discordance were 58.5% and 1.2%, respectively. Of major discordance, ER (48.2%, p< 0.001) and HER2 FISH (50.0%) led to more management changes than HER2 IHC (2.4%, p= 0.04) and PR (1.6%, p= 0.10). Patients with ER major discordance had increased risk of death (6.7% concordance vs. 22.2% major discordance, p= 0.004). Overall, retesting ER and HER2 was more clinically beneficial than retesting PR. To aid decision-making and minimize healthcare costs, we propose patient-centered guidelines on retesting biomarker profiles.

NOHA: A sensitive, low-cost, and accessible blood-based biomarker to determine breast cancer estrogen receptor status in low-resource settings.

580 Background: Significant challenges to breast cancer control in low- and middle-income countries (LMICs) include late-stage disease presentation because of few/no early detection programs, inadequately staffed and equipped pathology laboratories, and constrained treatment options. Estrogen receptor (ER) expression is critical to determining candidacy for cost efficient and accessible hormonal agents in LMICs; however, access to standard immunohistochemistry (IHC)-based ER analysis is grossly limited/nonexistent due to cost and technical requirements. We have identified Nw-hydroxy-L-Arginine (NOHA) as a low cost and accessible blood-based biomarker to distinguish estrogen-receptor negative (ER–) from estrogen-receptor positive (ER+) breast cancer, differentiate ER– high grade versus low grade tumors, and correlate ER– molecular phenotype with ethnic variation. Our studies with US patients suggest the NOHA threshold of &lt;4nM as a reliable indicator of ER– versus ER+ disease (Table 1). Here we examine the clinical utility of NOHA as an alternative to IHC in distinguishing ER– from ER+ breast tumors in Tanzanian patients. Methods: Following informed consent, 70 newly diagnosed breast cancer patients were recruited at Kilimanjaro Christian Medical Center (KCMC; Moshi, Tanzania). Prior to any treatment, a needle prick amount of blood was collected from each patient on a Noviplex plasma card (Shimadsu, U.S.) and stored at -80˚C. Plasma cards and unstained tumor pathology slides were shipped at 2-3 months intervals to US labs for NOHA and immunohistochemistry (IHC) ER testing. Statistical difference was set at p&lt;0.01, with NOHA and IHC assay operators blinded to patient clinical status. Results: Our early data show correlation between NOHA levels and ER IHC results, providing a means to distinguish ER– from ER+ breast cancer in the low-resource setting. Plasma cards stored at -80˚C for up to 3 months retained NOHA stability in assays involving a proprietary antibody-based ELISA, and by LC-MS. Conclusions: This study suggests the clinical utility of NOHA as a cost-effective, accessible replacement for standard IHC testing in determining ER status among breast cancer patients in LMICs, promising to extend access to cost efficient and available hormonal agents and improving outcomes and quality of life. The present study provides foundational knowledge for broader studies of NOHA utility in global breast cancer control, as well as in ongoing development of NOHA rapid-testing technologies. [Table: see text]