Estrogen Receptor Expression Research Articles

Estrogen promotes Epithelial ovarian cancer cells proliferation via down-regulating expression and activating phosphorylation of PTEN

Epithelial ovarian cancer (EOC) is the most common of cancer death among malignant tumors in women, its occurrence and development are strongly linked to estrogen. Having identified the phosphatase and tensin homologue (PTEN) is a potent tumor suppressor regulating cell proliferation, migration, and survival. Meanwhile, there is a correlation between PTEN protein expression and estrogen receptor expression in EOC. However, no study has amplified on the molecular regulatory mechanism and function between estrogen and PTEN in the development of EOC. In this research, we found that PTEN shows a low expression level in EOC tissues and estrogen decreased PTEN expression via the estrogen receptor 1 (ESR1) in EOC cells. Knockdown of PTEN enhanced the proliferation and migration level of EOC cells driven by estrogen. Moreover, PTEN was also phosphorylated by G protein-coupled receptor 30 (GPR30)-Protein kinase C (PKC) signaling pathway upon estrogen stimulation. Inhibiting the phosphorylation of PTEN weakened the proliferation and migration of estrogen induced-EOC cells estrogen and decreased the phosphorylation of Protein kinase B (AKT) and Mammalian target of rapamycin (mTOR). These results indicated that estrogen decreased PTEN expression level via the ESR1 genomic pathway and phosphorylated PTEN via the GPR30-PKC non-genomic pathway to activate the PI3K/AKT/mTOR signaling pathway, thereby determining the fate of EOC cells.

Primary Mucinous Cystadenocarcinoma of the Breast Intermixed with Pleomorphic Invasive Lobular Carcinoma: The First Report of This Rare Association.

Primary mucinous cystadenocarcinoma (MCA) is a rare breast carcinoma subtype showing overlapping histopathological features with mucinous cystadenocarcinoma of the ovary and pancreas. Current literature data suggest a favorable prognosis of breast MCAs despite its immunoprofile usually revealing lack of expression of estrogen receptor, progesterone receptor and HER-2 and high Ki67. As far as we know, only 36 cases have been reported in the literature to date. Its ambiguous morpho-phenotypic profile makes histological diagnosis very challenging. It must be distinguished from typical mucin-producing breast carcinomas and, above all, metastases from the same histotype in other sites (ovary, pancreas, appendix). Herein, we report the case of a primary breast MCA occurring in a 41-year-old female with peculiar histological features.

Negative estrogen receptor expression assessed by 18F-FES PET in metastatic breast cancer with ER-positive primary tumor.

1034 Background: The 18F-fluoroestradiol positron emission tomography/computed tomography (18F-FES PET/CT) technique provides a convenient method to evaluate the overall estrogen receptor (ER) expression in metastatic breast cancer (MBC) patients. There are long debates on the characteristics and treatment strategy of patients with positive primary ER lesion but negative ER expression in metastatic disease. 18F-FES PET offers an opportunity to answer this question. Methods: This study enrolled MBC patients with ER-positive primary tumor who received 18F-FES PET/CT in our center. Descriptive statistics were used in clinicopathologic characteristics and compared with Chi square test or t test. Progression‐free survival (PFS) was estimated by Kaplan-Meier method and compared by log-rank test. Results: 16.46% (52/314) patients with an ER-positive primary tumor had negative ER expression assessed by 18F-FES for MBC prior to receiving first-line systemic therapy. The rate of ER-negativity tested by 18F-FES was negatively co-related with levels of ER expression in early stage, with a positive to negative conversion in 2 of 3 (46.3%) ER-low (ie, ER 1%-9%) tumors, 7 of 10 (70.0%) ER-moderate (ie, ER 10%-49%) tumors, 16 of 114 (14.0%) ER-high (ie, ER 50%-95%) tumors, and 1 of 8 (12.5%) ER-very high (ie, ER > 95%) tumors (p < 0.001). Chemotherapy (83.33%, 40/48) was the most common treatment strategy afterward, among which capecitabine monotherapy (64.29%, 27/42) was a dominant alternative. PFS was significantly prolonged with capecitabine alone versus other chemotherapy (median PFS: 13.14 vs 6.21 months, p = 0.035). Conclusions: 18F-FES could identify negative conversion of ER in MBC which occurred frequently. Patients with lower ER expression in the primary lesion were more likely to have negative ER expression in the metastasis. In real-world clinical practice, chemotherapy was the primary choice by most physicians and capecitabine monotherapy had shown good efficacy.

Immunohistochemical expression of ER and p53 as predictive biomarkers of immunotherapy in endometrial cancer.

e17626 Background: Immune Checkpoint Inhibitors (ICIs), either as monotherapy or in combination with tyrosine kinase inhibitors (TKIs), were recently incorporated into the therapeutic guidelines for recurrent/metastatic endometrial cancer (EC). Microsatellite Instability (MSI) is the only biomarker to guide treatment. We evaluated the predictive significance of Estrogen Receptor (ER) and p53 expression. Methods: Medical charts of EC patients that received ICIs in our institution for advanced/metastatic disease were retrospectively reviewed. TP53 and ER expression were determined by immunohistochemistry. Survival analysis was performed using IBM SPSS statistics v.25. Results: Thirty-two patients were included in our analysis. The median age was 63.1 years (25th-75th percentile: 57.6 – 69.6). 62.5% of the patients had endometrioid histology, 21.9% serous histology and 15.6% of them displayed other histology. 46.9% of the patients had grade 1-2 tumors and 53.1% grade 3 tumors. 21.9% of them were diagnosed with stage 1-2 disease, while 78.1% presented with stage 3 disease. 62.5% received 0-1 prior lines of treatment and 37.5% of them received more than 2 prior lines. Molecular characteristics are presented in the table. All MSI-High patients received anti-PD1 monotherapy with a median PFS of 11.9 months (95% CI:2.3-21.5). Microsatellite Stable (MSS) patients received pembrolizumab-lenvatinib combination with median PFS 7.54 months (95% CI:2.6-11.5). ER positivity was associated with significant PFS benefit in both the ICI monotherapy group (11.9 vs. 0.63 months, p=0.061) and the group that received ICI/TKI combination (11.3 vs. 1.75 months, p=0.001). Analogous benefit was noted in the Overall Survival post start of IO or IO/TKI treatment (9.8 vs 0.6 months in MSI-high patients and 20.6 vs 5.9 months in MSS patients). ER-positivity remained an independent predictive factor favoring PFS in the Multivariate Cox regression analysis when adjusting for immunotherapy type, histology, and previous lines of therapy (HR=0.216, p=0.013). Conclusions: ER expression proved to be an independent prognostic factor regarding PFS in our cohort of patients. Further studies are needed to investigate the role of ER as a predictive biomarker for immunotherapy in EC. [Table: see text]

Vascular Endothelial Growth Factor Expression in Breast Carcinoma and its Association with Expression of Estrogen Receptors, Progesterone Receptors and HER-2/neu and Molecular Subtypes

Background: The management of breast carcinoma depends on several molecular markers and tumor stages. In the last decades, estrogen receptors (ER), progesterone receptors (PR), and HER-2/neu have shown good therapeutic responses. Among other molecular markers, vascular endothelial growth factor (VEGF) is becoming more widely used as a prognostic indicator in patients with breast carcinoma. Anti-VEGF therapy already has been proven as an effective chemotherapeutic agent in some other carcinomas. The study aimed to find out the immunohistochemical expression of Vascular Endothelial Growth Factor (VEGF) in breast carcinoma and its possible correlation with the expression of ER, PR, and HER-2/neu and molecular subtypes to evaluate its prognostic value. Material & Methods: This study was conducted in the Department of Pathology, BIRDEM General Hospital, Dhaka, from March 2018 to January 2020. In this study, 45 diagnosed cases of breast carcinoma were enrolled. Slides of all cases were stained with ER, PR, HER-2/neu, and VEGF antibodies following the avidin-biotin-peroxidase staining method. Results: Among 45 cases, 60% showed positive immunohistochemical expression of VEGF. Most of these cases (71.1%) were ER/PR positive. VEGF did not show a significant association with other molecular markers or molecular subtypes. Conclusion: Although, the potential prognostic value of VEGF has not been confirmed. Based on the findings of the current study, it can be assumed that VEGF plays an important role in the pathogenesis of breast cancer. So, it may serve as a useful biomarker for immuno-targeting therapy in patients with breast cancer.

The role of local expression of hormone receptors in the genesis of idiopathic tracheal stenosis.

Tracheal stenosis in adults is usually the result of mechanical injuries either from direct trauma, tracheotomy or intubation. Idiopathic stenosis in the cricotracheal section is a rare condition and occurs almost exclusively in females. Therefore, an influence of the female sexual hormones estrogen and progesterone has been assumed previously. Tracheal specimens of 27 patients who received tracheal resection for either idiopathic tracheal stenosis (ITS) (n=11) or posttraumatic tracheal stenosis (PTTS) (n=16) between 2008 and 2019 in our surgical department were included and retrospectively analyzed. Immunohistochemical staining of tracheal specimens concerning the hormone receptor status of progesterone and estrogen was performed. While post-tracheotomy stenosis occurred in males (n=6) as well as in females (n=10), none of the patients with idiopathic stenosis were males. All of the idiopathic stenosis (n=11; 100%) showed a strong expression of the estrogen receptors (ERs) in the fibroblasts and expression of progesterone receptors (PRs) in fibroblasts in 8 of 11 (72.7%). In the post-tracheotomy patients, only 3/16 (18.8%) showed slight staining of PRs and 6/16 (37.5%) of ERs. Of those, only one male patient presented with expression of ERs and PRs and another male patient presented with isolated PRs. Oral intake of hormone compounds was seen in 11/27 (40.7%) patients: 7/11 (63.6%) in the ITS group and 4/16 (25%) in the PTTS (noteworthy that the PTTS group included 6 male patients). Although the number of patients is small, our results show that the expression of female sexual hormone receptors in the fibroblasts of the trachea is a persistent finding in ITS. Surgery provided good results with a favorable long-term outcome without recurrence of stenosis for ITS and PTTS. Further investigation with a special focus on hormones is needed to assist in the prevention of this rare disease.

Brain Metastasis is Associated with Tumor Size, Nodal Status, and c-erbB-2 Expression in Invasive Breast Carcinoma

Abstract Introduction According to the World Health Organization (WHO) classification, invasive breast carcinoma (IBC) of no special type (IBC-NST) is the second most common primary site of central nervous system metastases, affecting 15% to 30% of patients. Brain metastasis originating from IBC is associated with patient age, tumor size, and axillary lymph node status. Loss of expression of hormone receptors and c-erbB-2 amplification are frequent findings in patients who develop brain metastasis. Radiological studies of the central nervous system are carried out only in patients presenting with neurological signs or symptoms during the clinical follow-up. Objective To evaluate the associations of clinical and pathological findings with brain metastasis in breast cancer. Materials and Methods The sample comprised 73 patients with breast cancer who underwent mastectomy with lymph node resection. The following variables were evaluated: tumor size, histological grade, nodal state, expression of estrogen and progesterone receptors and c-erbB-2, and presence of brain metastasis. Results The histopathological findings associated with brain metastasis in patients with IBC were tumor size (p = 0.03), presence of nodal metastasis (p = 0.045), and c-erbB-2 expression (p = 0.012). Conclusion The assessment of specific pathological findings in breast carcinoma can help identify risk factors and/or clinical parameters associated with the development of brain metastasis.

Antitumor activity of AX-0085, a novel AXL kinase inhibitor: Analysis of apoptotic cell death in triple-negative breast cancer cells.

e15120 Background: Triple-negative breast cancer (TNBC) is insensitive to targeted therapies due to the negative expression of human epidermal growth factor receptor-2, estrogen receptor and progesterone receptor. TNBC cells express higher levels of AXL more often than other breast cancer subtypes. The receptor tyrosine kinase AXL plays a role in survival, invasion, migration, epithelial-mesenchymal transition (EMT), metastasis, resistance, and immune suppression in cancer cells including in TNBC. Thus, AXL is a promising therapeutic target for the treatment of TNBC, and we developed a new small-molecule AXL inhibitor, AX-0085. Methods: The interaction mechanism between AX-0085 and AXL was studied by molecular dynamic simulations, cell-based kinase assay, and western blotting in TNBC cells. We investigated the anti-tumor activity of AX-0085 on the AXL-dependent pro-tumorigenic properties such as cell proliferation, invasion, migration, EMT, and immune suppression (PD-L1 expression) in TNBC cell lines and investigated in vivo efficacy of AX-0085 with the 4T1 xenograft model. Results: In the molecular dynamic simulations, estimated binding energy of AX-0085 and cabozantinib to AXL was -10.1Kcal/mol and -8.1Kcal/mol, respectively. AX-0085 was made a hydrogen bond with catalytically important residue Lys567 and blocked the activation of AXL by inhibiting the salt bridge with Lys567-Glu585. AX-0085 (IC50: 4.4nM for AXL kinase in cell-based kinase assay) was one of the most potent AXL inhibitors. AX-0085 showed a high antitumor activity against all TNBC subtypes and was a more potent anti-proliferative activity than other kinase inhibitors in MDA-MB-231 cell line. AX-0085 effectively blocked the activation of AXL in TNBC and further inhibited AXL-dependent pro-tumorigenic events such as cell proliferation, invasion, migration, EMT, and PD-L1 expression in TNBC cell. AX-0085 induced cell cycle arrest at G1 phase and suppressed the expression of CDK2 and Cyclin E in TNBC. AX-0085 promoted apoptotic cell death by up-regulating cleaved caspase-9 and down-regulating Bcl-2 in TNBC. Pharmacological investigation showed that the AX-0085 treated tumors displayed a dose-dependent reduction in volume and weight of the tumors in the 4T1 mouse xenograft. Conclusions: Our results demonstrated that AX-0085 selectively blocks AXL activation which confers an effective therapeutic value in the treatment of TNBC. Currently, AX-0085 is undergoing non-clinical trials.

Artificial intelligence (AI) –based machine learning models (ML) for predicting pathological complete response (pCR) in patients with hormone receptor (HoR) –positive/HER2-negative early breast cancer (EBC) undergoing neoadjuvant chemotherapy (NCT): A retrospective cohort study.

597 Background: The rates of pCR after NCT in HoR positive/HER2 negative EBC are low. Thus, clinicians need validated, unbiased, and reliable tools to better predict pCR at individual patients’ level. In this regard, AI can generate ML algorithms with these features, using clinical and pathological characteristics. Methods: Medical records were retrospectively retrieved for patients with EBC receiving NCT followed by surgery. Missing data were imputed using chained random forests. pCR was defined as absence of residual invasive cancer on pathologic evaluation of the breast specimen and lymph nodes (ypT0/isN0). Differences between pCR and non-pCR patients were assessed using t test or chi-squared test. Eight ML models (c5.0, k-nearest neighbour, random forest [RF], neural network, support vector machine [linear/radial], boosted trees and boosted logistic regression) were trained and tuned. A stratified ten-fold cross-validation was used to prevent overfitting. Models performance was evaluated using the area under the receiver operating characteristics curve (AUC). Features importance was assessed. Disease-free survival (DFS) was evaluated using the Kaplan-Maier method and hazard ratios (HRs) using Cox regression. Results: 572 patients were included: 332 (58%) were pre-menopausal and median age was 49.0 (IQR 43-57); 330 (58%) were T2 and 273 (48%) were N1; 437 (76%) had a ductal adenocarcinoma and 308 (54%) were grade 3. Median estrogen and progesterone receptor (ER/PgR) expression was 90 (IQR 80-90) and 57.5 (IQR 5-90); median Ki67 was 35 (IQR 25-55) and 230 (40%) patients were HER2 zero. 565 (99%) patients received a combination of anthracyclines and taxanes; 434 (76%) received sequential and 137 (24%) concomitant CT. pCR was achieved in 87 patients (15%, 95% CI 12-18). Ten variables were included in the model: menopausal status, age, histology, grade, clinical T and N stage, ER/PgR status, Ki67 and HER2 status (zero/low). Among the evaluated models, the RF algorithm had the best performance: the AUC was 0.77 (95% CI 0.71-0.83) and sensitivity and specificity were 0.86 (95% CI 0.82-0.88) and 0.56 (95% CI 0.46-0.66). Variables with the highest importance were Ki67, ER/PgR status, age and nodal status. 511 patients were evaluable for survival: patients with pCR had a significant longer DFS compared to those who did not achieve pCR (HR 0.30; 95% CI 0.14-0.65, p = 0.002). Patients whose pCR was predicted by the model had longer DFS compared to those who pCR was not predicted (HR 0.56, 95% CI 0.21-0.87, p = 0.01). Conclusions: The RF-ML algorithm combining clinical and pathological characteristics has the potential to predict pCR in HoR positive/HER2 negative patients undergoing NCT, thus supporting clinicians to individualize treatment for EBC.

Real-world characteristics and treatment pattern of patients with newly diagnosed endometrial cancer in China.

e17613 Background: Endometrial cancer (EC) is the second most common gynecologic cancer in China with an increasing incidence. Currently, there is no comprehensive profile of EC patients in China. This real-world multicenter retrospective study aimed to depict the characteristics and treatment patterns in newly diagnosed EC patients in China. Methods: Medical records of newly diagnosed EC at 8 hospitals between January 1, 2015 and December 31, 2019 were retrospectively reviewed. The demographic and clinical characteristics at diagnosis as well as the treatment received within 6 months after diagnosis were collected. Results: A total of 1397 patients were enrolled in the study. The average age was 53.5 years. The number of patients (n=1256) classified as stage I, II, III, and IV was 885 (70.5%), 141 (11.2%), 207 (16.5%) and 23 (1.8%), respectively. The number of patients (n=1340) presented with low-grade (G1-2) and high-grade (G3) tumors was 924 (74.0%) and 324 (26.0%), respectively. Invasion of lymphovascular space was detected in 237 (17.0%) out of 1397 patients. Among 1389 patients with pathological data, pure endometrioid carcinoma (90.3%) and serous carcinoma (5.1%) accounted for the majority of pathological types. Estrogen receptor (ER) expression was positive in 91.0% (977/1074) of patients and progesterone-receptor (PR) expression was positive in 85.2% (904/1061) of patients. Mismatch repair deficient/microsatellite instability-high (dMMR/MSI-H) was identified in 26.6% (107/402) of patients. Among 1393 patients with treatment information, surgery was performed in 99.6% of patients and 66.9% received adjuvant treatment following surgery according to clinicopathological risk factors. The percentage of patients receiving adjuvant chemotherapy, adjuvant radiotherapy, and adjuvant chemoradiotherapy was 36.7%, 7.3%, and 22.0%, respectively. Among 306 patients who received chemoradiotherapy, 47.1% underwent concurrent chemoradiotherapy, 31.0% underwent sandwich chemoradiotherapy, and 21.9% underwent sequential chemoradiotherapy. Conclusions: To date, this is the first real-world study with largest sample size of EC patients in China. EC is frequently diagnosed at early 50s in Chinese patients, which is at a younger age compared with European and American population. Newly diagnosed EC patients in China are primarily treated with surgery. Chemotherapy and radiotherapy remain the main adjuvant treatment after surgery. MSI-H/dMMR patients accounted for about 1/4 of newly diagnosed patients, for whom immunotherapy may open up better prognosis in the future.

Prognostic significance of estrogen receptor status in de novo metastatic breast cancer.

e13049 Background: The clinical behaviors of de novo metastatic breast cancer (dnMBC) differ according to hormone receptor (HR) status, but an association between estrogen receptor (ER) expression and clinical outcomes has not been established. Our study aimed to investigate the influence of different ER status on survival outcomes for patients with dnMBC. This study was the Medical Science Research Project of Hebei Province and approved by the Ethics Committee of the Ethics Committee of Cangzhou Clinical College of Integrated Traditional Chinese and Western Medicine of Hebei Medical University (NO. 2017-AF29-058). Methods: Using hospital-based database, we identified patients with dnMBC diagnosed between 2010 and 2017, retrospectively collecting demographic data, tumor characteristics, treatment types and survival data. According to ER status, the patients were divided into ER-negative and ER-positive groups, and the propensity score matching (PSM) was used to balance the mixed bias. Overall survival (OS) was estimated by Kaplan-Meier method and log rank test. Univariable and multivariable analyses were performed using the Cox proportional hazard model to identify statistically significant prognostic factors. Results: A total of 322 patients were included. The median age was 53 (21-82) years, and the median follow-up was 8.5 years. The median OS for the PSM cohort was 29 months (95% confidence interval [CI], 26-35 months), and 5-year OS was 18.9%. OS differed significantly between the two groups (p = 0.0012), with a median OS of 36 months (95% CI, 32-41 months) in ER-positive group and 24 months (95% CI, 19-27 months) in ER-negative group. Multivariate analysis showed that clinical T-stage and initial metastatic sites were independent prognostic factors for survival in patients focused on ER status, with Ki-67 index, endocrine therapy and radiotherapy added to the ER-positive group, while initial and advanced chemotherapy added to the ER-negative group. Conclusions: For patients with dnMBC, there were substantial differences in OS according to ER status. ER-negative subtype is clinically different from ER-positive subtype, showing unfavorable tumor characteristics and poorer OS. We identified several prognostic factors that could guide therapy selection in this patient population. Clinical trial information: 20210957 .

Early prediction of endocrine responsiveness in ER+/HER2 negative MBC: Pilot study with 18F-fluoroestradiol (18F-FES) CT/PET.

1024 Background: Estrogen receptors (ER) are considered predictive biomarkers to identify ER+ MBC patients who would likely benefit from endocrine therapies (ET). However, 30 to 40% of ER+ MBC patients fail to achieve a durable response. 18F-FES PET/CT has been shown to represent a valuable and accurate diagnostic tool to determine ER status at the level of metastatic sites and can be proposed as a valid alternative to biopsy of metastatic lesions. In this trial, we evaluated 18F-FES PET/CT as a predictive tool for endocrine responsiveness in ER+ MBC. Methods: ET-FES is an international, multicenter, academic clinical trial, conducted within the JTC-2011 ERA-NET TRANSCAN programme. Patients with ER+/HER2- MBC and first evidence of relapse were eligible for the study. All patients underwent a baseline 18F-FES PET/CT in addition to conventional staging procedures. Tumors were classified as endocrine sensitive if overall Standardized Uptake Value (SUV) ≥ 2; SUVmax was measured in the 3 largest lesions to quantify ER expression and a cutoff value of 2 was used to dichotomize results (endocrine sensitive vs resistant). In the ET-FES trial, patients with SUV ≥ 2 received single agent ET until PD; patients with SUV < 2 were randomized to receive single agent ET or chemotherapy (CT). The predictive role of 18F-FES PET/CT results was assessed for PFS and OS by univariate and multivariate analyses. Results: Overall, 147 patients (142 for ITT analysis) were enrolled from 04/2015 to 12/2020; 113 presented with 18F-FES SUV ≥ 2 and received ET; 29 pts, with SUV < 2 were randomly assigned to ET or first line chemotherapy (CT), following local clinical practice. After a median follow up of 4.6 years, 51 deaths had occurred (54.2%). Median PFS was 18,0 months (95%CI 11,2- 22,9) in the overall population and in 18F-FES SUV ≥ 2 versus 12,4 months (95%CI 3,1 – NR) in patients with SUV <2 treated with ET and 23.0 months (95%CI 7,7 – 30,0) if treated with CT. Median OS was not reached in the overall patient population and in patients with SUV ≥2, treated with single agent ET. Median OS was 28.1 months (95%CI 14,2 – NR) in patients with SUV <2 treated with ET versus 52,8 months (95%CI 16,1 – NR) if treated with CT. The Kaplan–Meier estimate of OS at 48 months was 64.4% (SE +/-5%) in all patients and 66% in 18F-FES SUV ≥ 2; at 60 months was 54.4% (SE +/-6%) and 57% (SE +/-5%), respectively. Among the 113 pts with SUV ≥ 2 treated with ET, the Kaplan–Meier estimate of OS at 60 months was 73.0% if treated with aromatase inhibitors versus 35% in case of fulvestrant or tamoxifen (p< 0.0005). Conclusions: ET-FES is the first prospective trial to assess the efficacy of ET alone in patients selected for endocrine responsiveness on the basis of whole body molecular ER imaging by 18F-FES PET/CT. These results suggest that the efficacy of ET may be maximized by using the appropriate assessment of endocrine responsiveness at the different metastatic sites. Clinical trial information: EUDRACT 2013-000-287-29 .

Prognostic role of HER2 expression in patients with ER-positive/HER2-negative breast cancer: Results from a population-based cancer registry study.

547 Background: Estrogen Receptor (ER)-positive (+)/Human Epidermal Growth Factor Receptor 2 (HER2)-negative (-) breast cancers (BCs) express variable protein levels of both ER and HER2, which can influence prognosis. Methods: We evaluated all invasive ER+/HER2- BCs (n = 3633) that were diagnosed and systematically collected by the Parma Province Cancer Registry, Italy, from 2004 to 2018. Tumors were classified by HER2 (IHC score of 0, 1+ or 2+ with negative FISH) and ER status (ER-low [1-9%], ER-moderate [10-79%] or ER-high [80-100%]). Comparisons of clinicopathologic characteristics and disease outcome were performed. Results: BCs with late-stage diagnosis ( P = 0.04), high histologic grade ( P < 0.0001), or high proliferative rate ( P < 0.0001) were more likely HER2 2+/FISH-. The rate of ER-high BCs did not change from 2675 of 2938 (91%) HER2 0 tumors to 508 of 560 (90.7%) HER2 1+, and 124 of 135 (91.9%) HER2 2+/FISH- tumors. Correspondingly, ER-low BCs were not enriched among HER2 0 tumors compared to the other tumors with different HER2 expression ( P = 0.6). The 5-year overall survival (OS) for HER2 2+/FISH- BCs was lower than that for HER2 0 or 1+ tumors ( P = 0.03). ER-low/moderate tumors were associated with poorer OS in comparison with ER-high BCs ( P < 0.0001). HER2 2+/FISH- status was detrimental to OS among patients (pts) with ER-high tumors ( P = 0.04), while this finding was not observed among ER-low/moderate BCs ( P = 0.21). An interaction between HER2 2+/FISH- expression and ER-high status was found for poorer OS after adjusting for prognostic variables (HR = 1.7; 95% CI: 1.1-2.9). Conclusions: The prognostic role of HER2 expression in pts with ER-positive/HER2-negative BCs seems to be restricted to ER-high tumors, while the worse prognosis of tumors with lower ER expression is not associated with HER2 status. These findings may help identify optimal patient inclusion criteria for clinical trials with novel anti-HER2 therapies in ER-positive/HER2-negative disease.

Understanding the Phytoestrogen Genistein Actions on Breast Cancer: Insights on Estrogen Receptor Equivalence, Pleiotropic Essence and Emerging Paradigms in Bioavailability Modulation.

Prevalent as a major phenolic ingredient of soy and soy products, genistein is recognized as an eminent phytoestrogen owing to its interacting ability with estrogen receptors (ERs). The metabolic conversion of plant-derived genistin to genistein by gut microbes and intestinal enzymes enhances its absorption at intestinal pH of ~7.5-7.8. Genistein interferes in breast cancer (BC) development via pleiotropic actions on cell proliferation, survival, angiogenesis, and apoptosis. Though multiple investigations have demonstrated genistein intake-driven reduced BC risk, similar efficacy has not been replicated in clinical trials. Furthermore, multiple studies have structurally and functionally equated genistein extents with 17-β-estradiol (E2), the most available physiological estrogen in females, culminating in aggravated BC growth. Of note, both genistein and E2 function via interacting with ERs (ERα and ERβ). However, although E2 shows almost equal affinity towards both ERα and ERβ, genistein shows more affinity towards ERβ than ERα. Our cautious literature survey revealed typical intake mode, ER expression pattern and the ratio of ERα and ERβ, transactivators/ regulators of ERα and ERβ expression and activities, patient age, and menopausal status as decisive factors affecting genistein BC activities. Of further interest are the mechanisms by which genistein inhibits triple-negative breast cancers (TNBCs), which lack ERs, progesterone receptors (PRs), and human epidermal growth factor receptors (HER2). Herein, we attempt to understand the dosage-specific genistein actions in BC cells and patients with an insight into its better response via derivative development, nanocarrier-assisted, and combinatorial delivery with chemotherapeutic drugs.

Association of mutation and expression of the brother of the regulator of imprinted sites (BORIS) gene with breast cancer progression.

The BORIS, 11 zinc-finger transcription factors, is a member of the cancer-testis antigen (CTA) family. It is mapped to chromosome number 20q13.2 and this region is genetically linked to the early onset of breast cancer. The current study analyzed the correlation between BORIS mutations and the expression of the protein in breast cancer cases. A population-based study including a total of 155 breast cancer tissue samples and an equal number of normal adjacent tissues from Indian female breast cancer patients was carried out. Mutations of the BORIS gene were detected by polymerase chain reaction-single standard confirmation polymorphisms (PCR-SSCP) and automated DNA sequencing and by immunohistochemistry for BORIS protein expression were performed. The observed findings were correlated with several clinicopathological parameters to find out the clinical relevance of associations. Of all the cases 16.12% (25/155) showed mutations in the BORIS gene. The observed mutations present on codon 329 are missense, leading to Val> Ile (G>A) change on exon 5 of the BORIS gene. A significant association was observed between mutations of the BORIS gene and some clinicopathological features like nodal status (p = 0.013), estrogen receptor (ER) expression (p = 0.008), progesterone receptor (PR) expression (p = 0.039), clinical stage (p = 0.010) and menopausal status (p = 0.023). The protein expression analysis showed 20.64% (32/155) samples showing low or no expression (+), 34.19% (53/155) with moderate expression (++), and 45.17% (70/155) showing high expression (+++) of BORIS protein. A significant association was observed between the expression of BORIS protein and clinicopathological features like clinical stage (p = 0.013), nodal status (p = 0.049), ER expression (p = 0.039), and PR expression (p = 0.027). When mutation and protein expression were correlated in combination with clinicopathological parameters a significant association was observed in the category of high (+++) level of BORIS protein expression (p = 0.017). The BORIS mutations and high protein expression occur frequently in carcinoma of the breast suggesting their association with the onset and progression of breast carcinoma. Further, the BORIS has the potential to be used as a biomarker.

Effect of HER2 expression status on the prognosis of patients with HR+/HER2‑ advanced breast cancer undergoing advanced first‑line endocrine therapy.

The present study aimed to retrospectively assess the effects of human epidermal growth factor receptor 2 (HER2) expression on the diagnosis of patients with hormone receptor (HR)+/HER2- late-stage breast cancer undergoing advanced first-line endocrine-based treatment. A total of 72 late-stage breast tumor cases from June 2017 to June 2019 were selected from the Department of Surgical Oncology, Shaanxi Provincial People's Hospital (Xi'an, China) and included in the present study. The expression of estrogen receptor, progesterone receptor and HER2 was detected by immunohistochemistry. The subjects were divided into two groups: the HER2-negative (0) cohort (n=31) and the HER2 low expression cohort (n=41). The age, BMI, Karnofsky Performance Status (KPS) score, tumor size, lymph node metastasis, pathological type, Ki-67 expression and menopausal status of the patients were obtained through the electronic medical record system of Shaanxi Provincial People's Hospital. Progression-free survival (PFS) and overall survival (OS) were evaluated for all patients. The median PFS and OS of the HER2(0) cohort were longer than those of the HER2 low expression cohort (all P<0.05). It was shown that age (hazard ratio, 6.000 and 5.465), KPS score (hazard ratio, 4.000 and 3.865), lymph node metastasis (hazard ratio, 3.143; 2.983) and HER2 status (hazard ratio, 3.167 and 2.996) were independent influencing factors of the prognosis of patients with HR+/HER2- advanced breast cancer (ABC) (all P<0.05). Three models (model 1, no parameters adjusted; model 2, BMI, tumor size, pathological type, Ki-67 and menopausal status adjusted; and model 3, age, KPS functional status score and lymph node metastasis adjusted based on model 2) were established within the HER2(0) cohort as the reference for statistical analysis using the multivariate Cox's regression test. In models 2 and 3, the risk of poor prognosis of ABC within the HER2 low expression cohort was significantly higher compared with that in the HER2(0) cohort (hazard ratio, 3.558 and 4.477; 95% CI, 1.349-9.996 and 1.933-11.586; P=0.003 and P<0.001). The HER2 expression status of patients with HR+/HER2- ABC receiving advanced first-line endocrine therapy may affect PFS and OS.

Tumor-Infiltrating Lymphocytes and PD-L1 Expression in Pleomorphic Lobular Breast Carcinoma.

The prognostic and predictive role of stromal tumor-infiltrating lymphocytes (sTILs) is undetermined in pleomorphic invasive lobular cancer (pILC). The same applies for the expression of PD-1/PD-L1 in this rare breast cancer subtype. Here, we aimed to investigate the expression of sTILs and analyze the PD-L1 expression levels in pILC. Archival tissues from sixty-six patients with pILC were collected. The sTIL density was scored as a percentage of tumor area using the following cut-offs: 0%; <5%; 5-9%; and 10-50%. The PD-L1 expression was analyzed using IHC on formalin-fixed, paraffin-embedded tissue sections using SP142 and 22C3 antibodies. A total of 82% of the sixty-six patients were hormone receptor positive and 8% of cases were triple negative (TN), while 10% showed human epidermal growth factor receptor 2 (HER2) amplification. sTILs (≥1%) were present in 64% of the study population. Using the SP142 antibody, 36% of tumors demonstrated a positive PD-L1 score of ≥1%, and using the 22C3 antibody, 28% had a positive PD-L1 score of ≥1. There was no correlation between sTILs or PD-L1 expression and tumor size, tumor grade, nodal status, expression of estrogen receptor (ER), or amplification of HER2. Our data did not show any difference in survival between the three molecular subtypes of pILC with respect to sTILs and PD-L1 expression. This study shows that pILCs show some degree of sTILs and PD-L1 expression; however, this was not associated with a survival improvement. Additional large trials are needed to understand immune infiltration in lobular cancer, especially in the pleomorphic subtype.

Tissue-specific promoters regulate the transcription of cyp19a1 in the brain-pituitary-gonad axis of Japanese eel (Anguilla japonica)

Aromatase is a key enzyme that catalyzes the biosynthesis of estrogens. Previous study indicated that putative tissue-specific promoters of the one aromatase gene (cyp19a1) may drive the differential regulatory mechanisms of cyp19a1 expression in Anguilla japonica. In the present study, for elucidating the transcription characteristics and the function of putative tissue-specific promoters of cyp19a1 in the brain-pituitary-gonad (BPG) axis during vitellogenesis, we investigated the transcriptional regulation of cyp19a1 by 17β-estrogen (E2), testosterone (T), or human chorionic gonadotropin (HCG) in A. japonica. The expression of estrogen receptor (esra), androgen receptor (ara), or luteinizing hormone receptor (lhr) was up-regulated as cyp19a1 in response to E2, T, or HCG, respectively in the telencephalon, diencephalon, and pituitary. The expression of cyp19a1 was also upregulated in the ovary by HCG or T in a dose-dependent manner. Unlike in the brain and pituitary, the expression of esra and lhr, rather than ara, was upregulated by T in the ovary. Subsequently, four primary subtypes of 5′-untranslated terminal regions of cyp19a1 transcripts and the corresponding two 5′ flanking regions (promoter P.I and P.II) were identified. The P.II existed in all BPG axis tissues, whereas the P.I with strong transcriptional activity was brain- and pituitary-specific. Furthermore, the transcriptional activity of promoters, the core promoter region, and the three putative hormone receptor response elements were validated. The transcriptional activity did not change when the HEK291T cells co-transfected with P.II and ar vector were exposed to T. These results suggested that the expression of cyp19a1 was upregulated indirectly through esra and lhr rather than ara by T in the ovary, whereas the expression of cyp19a1 was upregulated directly through androgen receptor and the downstream androgen response element of tissue-specific P.I in the brain and pituitary. The results of the study reveal the regulatory mechanisms of estrogen biosynthesis and provide a reference for optimizing the technology of artificially induced maturation in eels.

Glucocorticoid Receptors Drive Breast Cancer Cell Migration and Metabolic Reprogramming via PDK4.

Corticosteroids act on the glucocorticoid receptor (GR; NR3C1) to resolve inflammation and are routinely prescribed to breast cancer patients undergoing chemotherapy treatment to alleviate side effects. Triple-negative breast cancers (TNBCs) account for 15% to 20% of diagnoses and lack expression of estrogen and progesterone receptors as well as amplified HER2, but they often express high GR levels. GR is a mediator of TNBC progression to advanced metastatic disease; however, the mechanisms underpinning this transition to more aggressive behavior remain elusive. We previously showed that tissue/cellular stress (hypoxia, chemotherapies) as well as factors in the tumor microenvironment (transforming growth factor β [TGF-β], hepatocyte growth factor [HGF]) activate p38 mitogen-activated protein kinase (MAPK), which phosphorylates GR on Ser134. In the absence of ligand, pSer134-GR further upregulates genes important for responses to cellular stress, including key components of the p38 MAPK pathway. Herein, we show that pSer134-GR is required for TNBC metastatic colonization to the lungs of female mice. To understand the mechanisms of pSer134-GR action in the presence of GR agonists, we examined glucocorticoid-driven transcriptomes in CRISPR knock-in models of TNBC cells expressing wild-type or phospho-mutant (S134A) GR. We identified dexamethasone- and pSer134-GR-dependent regulation of specific gene sets controlling TNBC migration (NEDD9, CSF1, RUNX3) and metabolic adaptation (PDK4, PGK1, PFKFB4). TNBC cells harboring S134A-GR displayed metabolic reprogramming that was phenocopied by pyruvate dehydrogenase kinase 4 (PDK4) knockdown. PDK4 knockdown or chemical inhibition also blocked cancer cell migration. Our results reveal a convergence of GR agonists (ie, host stress) with cellular stress signaling whereby pSer134-GR critically regulates TNBC metabolism, an exploitable target for the treatment of this deadly disease.

(104) Immunohistochemical Staining Reveals Estrogen, Androgen and Progesterone Receptors in the Vestibule

Abstract Introduction The introitus has three embryologic tissue types: ectoderm (vulva), endoderm (vestibule), and mesoderm (vagina). The vulva and vagina have been shown previously to have estrogen receptors (ER), androgen receptors (AR) and progesterone receptors (PR). ER and AR have been demonstrated in vaginal epithelial cells, as well as in fibroblasts and smooth muscle of underlying stroma. Vulvar ER and AR were demonstrated in epidermal keratinocytes and dermal fibroblasts of hair and non-hair bearing skin. AR were also seen in vulvar sebaceous glands, sweat glands, and hair follicles. PR was seen in vaginal epithelium, fibroblasts, and smooth muscle but not in the vulva. Despite the importance of the vestibule to sexual health, there is limited information regarding vestibular hormone receptors. Objective To better understand vestibular hormonal physiology, immunohistochemical (IHC) studies of ER, AR, and PR was performed on excised vestibulectomy tissue specimen. Methods IHC was performed on 7 vestibular specimen from eugonadal individuals with confirmed diagnosis of neuroproliferative vestibulodynia based on positive CD117 and PGP9.5 staining, consistent with excess mast cells and nerve endings, respectively. Specimen were also studied with primary antibodies (Anti-ERα, Ab 32063; Anti-AR, MI 06680; Anti-PR, Dako PGR636). Finally, sections were lightly counterstained with Gill-modified hematoxylin. Results ER and AR were richly expressed in tissue samples from all 7 patients. Dense labeling of both receptor types (AR on left, on ER right) was identified in the germinal layer of the epithelium (Fig. 1) with comparatively sparse expression in the subepithelial stroma. AR, but not ER, were noted in mucinous vestibular glands (Fig. 2). While labeling of PR was highly variable (Fig. 3), all samples showed evidence of moderate PR labeling in the subepithelial stroma. Variability may be due to menstrual cycle phase during which samples were obtained, however this information was not collected. Conclusions This preliminary study provides evidence of dense ER and AR expression in the vestibule, consistent with it being in part an androgenic organ. This supports the concept that the vestibule may be more effectively treated with combined testosterone and estradiol cream than with topical estradiol alone in patients with entrance dyspareunia from hormonally mediated vestibulodynia or genitourinary syndrome of menopause. Also consistent with these findings is that treatment with intravaginal DHEA, which converts to testosterone and estradiol intracrinologically and spreads to the vestibule, successfully resolves entrance dyspareunia, as shown in a previous clinical trial. While the significance of variable vestibular progesterone receptor expression is unclear, the addition of topical progesterone in patients with entrance dyspareunia may be considered. Disclosure Any of the authors act as a consultant, employee or shareholder of an industry for: Endoceutics, Inc