Loss Of Estrogen Receptor Research Articles

TRPS1 is a Highly Sensitive Marker for Breast Cancer: A Tissue Microarray Study Evaluating More Than 19,000 Tumors From 152 Different Tumor Entities.

Trichorhinophalangeal syndrome 1 (TRPS1) is a nuclear protein highly expressed in breast epithelial cells. TRPS1 immunohistochemistry (IHC) has been suggested as a breast cancer marker. To determine the diagnostic and prognostic utility of TRPS1 IHC, tissue microarrays containing 19,201 samples from 152 different tumor types and subtypes were analyzed. GATA3 IHC was performed in a previous study. TRPS1 staining was seen in 86 of 152 tumor categories with 36 containing at least one strongly positive case. TRPS1 staining predominated in various types of breast carcinomas (51%-100%), soft tissue tumors (up to 100%), salivary gland tumors (up to 46%), squamous cell carcinomas (up to 35%), and gynecological cancers (up to 40%). TRPS1 positivity occurred in 1.8% of 1083 urothelial neoplasms. In invasive breast carcinoma of no special type, low TRPS1 expression was linked to high grade ( P = 0.0547), high pT ( P < 0.0001), nodal metastasis ( P = 0.0571), loss of estrogen receptor and progesterone receptor expression ( P < 0.0001 each), and triple-negative status ( P < 0.0001) but was unrelated to patient survival ( P = 0.8016). In squamous cell carcinomas from 11 different sites, low TRPS1 expression was unrelated to tumor phenotype. Positivity for both TRPS1 and GATA3 occurred in 47.4% to 100% of breast cancers, up to 30% of salivary gland tumors, and 29 (0.3%) of 9835 tumors from 134 other cancer entities. TRPS1 IHC has high utility for the identification of cancers of breast (or salivary gland) origin, especially in combination with GATA3. The virtual absence of TRPS1 positivity in urothelial neoplasms is useful for the distinction of GATA3-positive urothelial carcinoma from breast cancer.

Implication of IZUMO2 in the cell-in-cell phenomenon: A potential therapeutic target for triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is characterized by the loss of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. The aggressive clinicopathological features and resistance to currently available therapeutics of the disease warrant an urgent need for the development of novel alternate therapeutic options. We have previously reported adiponectin-expressing regulatory T cells (A-Tregs), which can induce apoptosis in TNBC through the cell-in-cell phenomenon. In this study, we aimed to elucidate the molecule that allows TNBC cells to engulf A-Tregs. A monoclonal antibody, which repressed the engulfment of A-Tregs by TNBC cells, was developed. Immunoprecipitation followed by mass spectrometry and small interfering RNAs-mediated gene silencing was performed to characterize the antigen. We successfully generated a monoclonal antibody, designated G1D7, which abrogated the engulfment of A-Tregs by TNBC and subsequent A-Treg-mediated apoptosis. G1D7 detected the immunoglobulin-like type I membrane protein IZUMO2, a molecule related to IZUMO1 that is essential for cell-cell membrane binding and fusion of sperm to oocyte. The findings highlight the importance of IZUMO2 on TNBC cells in facilitating the cell-in-cell phenomenon by A-Tregs.

Clinicopathological features and survival outcomes of luminal-like breast tumors with estrogen receptor loss at metastatic recurrence: A case-control study

IntroductionEstrogen receptor (ER) loss at metastatic relapse occurs in up to 20% of luminal-like primary breast tumors. Data about clinicopathological features associated with ER loss and its prognostic significance are limited. MethodsIn a nested-case-control study, we compared clinicopathological characteristics and clinical outcomes between a cohort of 51 patients with primary ER+ /HER2- and paired triple-negative metastasis (LUM-TN) and two control cohorts of paired early-metastatic ER+ /HER2- (LUM-LUM, n = 50) and triple-negative (TN-TN, n = 49) breast cancers. Stromal tumor-infiltrating lymphocytes (TILs) were assessed according to the TILs Working Group recommendations as continuous and discrete variables with cutoffs (20%, 40%). ResultsLUM-TN tumors had lower ER expression than LUM-LUM tumors, but lower grade and Ki67 than TN-TN cases. Median distant-metastasis free survival was similar for LUM-TN and LUM-LUM cohorts, but significantly longer than in TN-TN cases (log-rank P < 0.001). LUM-TN and TN-TN cohorts had a comparable survival from the time of metastatic recurrence, which was significantly shorter than in patients with LUM-LUM tumors (log-rank P < 0.001). High TILs were associated with worse outcomes in patients with ER loss (P < 0.001). ConclusionsBreast tumors with ER loss at metastatic relapse have intermediate features and outcomes compared with metastatic luminal-like and ab initio triple-negative tumors. Further investigation on the biological mechanisms underpinning the loss of ER expression is ongoing.

Serine starvation silences estrogen receptor signaling through histone hypoacetylation

Loss of estrogen receptor (ER) pathway activity promotes breast cancer progression, yet how this occurs remains poorly understood. Here, we show that serine starvation, a metabolic stress often found in breast cancer, represses estrogen receptor alpha (ERα) signaling by reprogramming glucose metabolism and epigenetics. Using isotope tracing and time-resolved metabolomic analyses, we demonstrate that serine is required to maintain glucose flux through glycolysis and the TCA cycle to support acetyl-CoA generation for histone acetylation. Consequently, limiting serine depletes histone H3 lysine 27 acetylation (H3K27ac), particularly at the promoter region of ER pathway genes including the gene encoding ERα, ESR1. Mechanistically, serine starvation impairs acetyl-CoA-dependent gene expression by inhibiting the entry of glycolytic carbon into the TCA cycle and down-regulating the mitochondrial citrate exporter SLC25A1, a critical enzyme in the production of nucleocytosolic acetyl-CoA from glucose. Consistent with this model, total H3K27ac and ERα expression are suppressed by SLC25A1 inhibition and restored by acetate, an alternate source of acetyl-CoA, in serine-free conditions. We thus uncover an unexpected role for serine in sustaining ER signaling through the regulation of acetyl-CoA metabolism.

Abstract 5315: Identification of urinary proteome signatures associated with triple-negative breast cancer

Abstract Triple-negative breast cancer (TNBC), an aggressive breast cancer subtype, accounts for ~15% of all breast cancers and is associated with a poor prognosis and high rates of recurrence and distant metastases. Patients diagnosed with TNBC have limited treatment options and are often restricted to chemotherapy-based regimens, in stark contrast to patients with other breast cancer subtypes where the development of targeted therapeutics have significantly improved patient outcomes. TNBC is characterized by the loss of estrogen receptor, progesterone receptor and human epidermal growth factor receptor 2. This and the genetic and molecular tumor heterogeneity of TNBC has not only limited advances in the development of targeted therapy but also in the identification of biomarkers that can predict disease status and monitor treatment response. Advances in next-generation sequencing technologies have led to the identification of promising clinically actionable mutations however these tests often require invasive procedures to obtain tumor tissue and lack the ability to monitor treatment efficacy or detect recurrence, highlighting the need for non-invasive markers. A number of serum protein biomarkers have been evaluated in TNBC however none have yet to be successfully deployed in the clinic as indicators of disease. We have previously established that the urinary proteome can serve as a source of highly specific and sensitive biomarkers that can detect disease status and stage of multiple cancer types. Within the context of our global proteomics approach to cancer biomarker discovery, we have utilized the SomaScan discovery platform to identify 281 proteins that are differentially expressed in the urine of patients with TNBC compared to the urine of age- and sex-matched controls. The gene expression profiles of these proteins in TNBC tumor tissue were compared to those of normal breast tissue utilizing the R2 Genomics Platform. Proteins whose gene expression profile was consistent with the proteomic data were validated for the ability to distinguish between TNBC and non-TNBC diagnoses using mono-specific ELISAs of patient urine samples. These results demonstrate that a global proteomics approach coupled with clinical gene expression data can be effectively utilized to identify potential biomarkers for TNBC. These findings identify clinically relevant proteins that should be further explored as potential biomarkers in the development of non-invasive diagnostics for TNBC. (This work was supported by the Breast Cancer Research Foundation and the Nile Albright Research Foundation). Citation Format: Michael N. Lombardo, Roopali Roy, Susan Pories, Meg Lotz, Rama Aldakhlallah, Simon T. Dillon, Towia A. Libermann, Marsha A. Moses. Identification of urinary proteome signatures associated with triple-negative breast cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5315.

Liver X receptors and estrogen receptor β, two players in a rare subtype of NSCLC.

Liver X receptors (LXRαβ) play essential roles in the maintenance of the normal functions of macrophages, in modulation of immune system responses and cholesterol homeostasis. We have reported that LXRαβ-/- mice develop squamous cell lung cancer. We now report that those LXRαβ-/- mice, which live to 18-months of age, spontaneously develop a second type of lung cancer resembling a rare subtype of NSCLC (TTF-1 and P63-positive). The lesions are characterized as follows: a high proliferation rate; a marked accumulation of abnormal macrophages; an increase in the number of regulatory T cells; a remarkably low level of CD8+ cytotoxic T lymphocytes; enhanced TGFβ signaling; an increased expression of matrix metalloproteinases accompanied by degradation of lung collagen; and a loss of estrogen receptor β (ERβ). Because NSCLC is associated with cigarette smoking, we investigated the possible links between loss of LXRαβ and CS. A Kaplan-Meier Plotter database revealed reduced expression of LXRαβ and ERβ was correlated with low overall survival (OS). Thus, reduction of LXRαβ expression by cigarette smoking may be one mechanism through which CS causes lung cancer. The possibility that maintenance of LXRαβ and ERβ signaling could be used in the treatment of NSCLC needs further investigation.

Abstract 1776: Loss of estrogen receptor alters drug responsiveness and supports a basal-like phenotype in endocrine-resistant HER2+/ER+ breast cancer

Abstract At least 50% of human epidermal growth factor 2 (HER2) enriched breast cancer (BC) is estrogen receptor positive (ER+). In clinical trials, patients with HER2+ BC expressing ER respond poorly to neoadjuvant anti-HER2 therapy versus those lacking ER expression. Additionally, combining anti-HER2 with endocrine therapy (ET), like aromatase inhibitor (AI), does not significantly improve pathological complete response in HER2+/ER+ patients. In advanced or de novo HER2+/ER+ metastatic BC (MBC), treatment with ET plus anti-HER2 yields the highest 5-year overall survival. Unfortunately, in the real-world setting outside of clinical trials, just 23% of patients receive this combination, and nearly 40% receive ET alone. The clinical challenge associated with treating advanced HER2+/ER+ BC demands a better understanding of ER signaling and downstream pathways in HER2-driven BC. Our goal is to identify alternative targeted therapies and/or tailor therapeutic combinatorial strategies to treat and/or delay progression of HER2+/ER+ BC. In this study, we established two long-term estrogen (E2) deprived (LTED) HER2+/ER+ cell line models from BT-474 (BT) and MDA-MB-361 (MM) to mimic endocrine therapy resistance (ETR) to AI, and characterized the response of these cell lines to anti-HER2 and other ETs. We also performed whole-genome sequencing (WGS) and single-cell RNA sequencing (scRNAseq) to identify differences between parental and derived ETR-LTEDs. The ER expression was retained in BT-LTEDs but lost in MM-LTEDs as compared to parental cells. Additionally, ER transcriptional activity was confirmed by increased expression of ER-target genes upon E2 stimulation only in BT-LTEDs. However, both LTED variants were less responsive to fulvestrant and showed upregulation of pro-survival AKT signaling. Focusing on the now ER- MM-LTED model, WGS and targeted duplex sequencing identified a significant increase in exonic missense mutations, notably C&amp;gt;T and C&amp;gt;A. Several of the mutated genes encode for transcription and chromatin regulatory factors. scRNAseq analysis showed that MM-LTEDs shift from a luminal- to more basal-like phenotype, with enrichment of genes that regulate immune response and cell motility. Thus, loss of ER expression in MM-LTEDs on the mRNA and protein levels may explain observed intrinsic resistance to fulvestrant and gain of the basal-like phenotype. In summary, we report that our ETR BT- and MM-LTED models capture distinct ER-associated phenotypes of HER2+/ER+ BC. Additional studies are necessary to understand the functional significance of these missense mutations on the development of drug resistance in HER2+/ER+ BC. Ongoing studies are testing pharmacological inhibition of key upregulated genes associated with the basal phenotype to potentially delay disease progression and provide better treatments for ETR HER2+/ER+ BC. Citation Format: Shaymaa Bahnassy, Hillary Stires, Lu Jin, Stanley Tam, Yasmin J. Dunn, Brendan F. Kohrn, Lawrence A. Loeb, Manasi Balachandran, Mircea Podar, Matthew D. McCoy, Robert A. Beckman, Rebecca B. Riggins. Loss of estrogen receptor alters drug responsiveness and supports a basal-like phenotype in endocrine-resistant HER2+/ER+ breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1776.

Abstract P1-17-05: Benefit of endocrine therapy for metastatic breast cancer after loss of hormone receptor expression in patients with HR+/HER2- primary tumors

Abstract Background. Loss of estrogen receptor (ER) and/or progesterone receptor (PR) expression between primary breast tumors and metastases occurs in one-fourth of patients. Loss of hormone receptor (HR) expression is associated with inferior clinical outcomes however optimal treatment strategies in this setting have not been defined. Methods. Patients with HR+/HER2- primary breast tumors and subsequent HR-/HER2- metastatic biopsies, as determined by immunohistochemical staining, were identified using the University of North Carolina Metastatic Breast Cancer Clinical Database which includes patients diagnosed with metastatic breast cancer from 2001-2019. Patients with second primary breast cancers were excluded. Clinical features were assessed using chi-square testing. Survival was analyzed using univariate and multivariate Cox proportional hazards modeling. Results. 1,240 patients had known primary breast and metastatic biopsy receptor status. Of these, 427 patients had HR+/HER2- primary tumors with concordant metastases, while 71 patients had HR+/HER2- primary tumors with discordant metastases due to HR loss. Loss of HR expression was associated with higher grade (grade 3: discordant 59% vs concordant 38%, p=0.002), lower ER expression (ER ≥50%: discordant 56.8% vs concordant 93.1%, p&amp;lt;0.001) and less frequent dual ER/PR expression in the primary tumor (ER+/PR+: discordant 57.7% vs concordant 80.4%, p&amp;lt;0.001), as well as with shorter median disease-free interval (discordant 40.8m vs concordant 66.1m, p&amp;lt;0.001). Patients with HR loss were more likely to have brain, skin and distant lymph node metastases than patients with HR+ metastases. Of patients with HR loss, 54/71 patients (76.1%) had HR- biopsies at the time of metastatic diagnosis, prior to treatment in the metastatic setting. Six patients had de novo metastatic disease with synchronous HR+ primary tumors and HR- metastases without prior therapy in any setting. Two patients had HR- metastases at metastatic diagnosis with subsequent HR+ metastases consistent with the primary tumor while two additional patients had HR+ and HR- biopsies from distinct metastatic sites taken within two months of each other. Six patients had HR+ metastases at diagnosis and subsequent HR- metastases. A greater proportion of patients with HR loss received chemotherapy in the first-line metastatic setting (71.4% vs 24.8%, p&amp;lt;0.001). Median progression free survival on any first-line therapy was shorter for patients with HR loss (6.4m vs 11.2m, hazard ratio 1.8 (95% CI 1.18-2.74)). Median overall survival from initial diagnosis and metastatic diagnosis were also significantly shorter for patients with HR loss (70.4m vs 123.8m, hazard ratio 1.9 (95% CI 1.42-2.59) and 18.6m vs 45.9m, hazard ratio 2.0 (95% CI 1.49-2.73)). Of patients with HR loss at metastatic diagnosis, those who received endocrine therapy in the metastatic setting had superior outcomes compared to those who did not receive endocrine therapy (21.9m vs 9.6m, hazard ratio 0.303 (95% CI 0.14-0.65)). In multivariate analysis HR loss and lack of dual ER/PR expression on the primary tumor remained associated with inferior survival beyond metastatic diagnosis when accounting for age, race, T stage, nodal status and grade. Conclusion. Discordant HR status in untreated, synchronous breast primary tumors and metastases in patients with de novo metastatic breast cancer, as well as in patients with discordant metastatic biopsies obtained within two months of concordant biopsies, suggests tumor heterogeneity within individual patients irrespective of prior therapy. Improved outcomes with endocrine therapy in the metastatic setting despite loss of HR on the initial metastatic biopsy highlights the continued value of the primary tumor phenotype to guide therapy in the metastatic setting. Citation Format: Danielle File, Yara Abdou, Amy Wheless, Claire Dees, Lisa Carey. Benefit of endocrine therapy for metastatic breast cancer after loss of hormone receptor expression in patients with HR+/HER2- primary tumors [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P1-17-05.

Diagnostic and prognostic impact of cytokeratin 19 expression analysis in human tumors: a tissue microarray study of 13,172 tumors

To evaluate cytokeratin 19 (CK19) expression in normal and cancerous tissues, 15,977 samples from 122 tumor types and 608 samples of 76 normal tissue types were analyzed by immunohistochemistry (IHC). In normal tissues, CK19 expression occurred in epithelial cells of most glandular organs but was strictly limited to the basal cell layer of nonkeratinizing squamous epithelium and absent in the skin. CK19 expression in ≥90% of cases was seen in 34% of the tumor entities including the adenocarcinomas of the pancreas (99.4%), colorectum (99.8%), esophagus (98.7%), and stomach (97.7%), as well as breast cancer (90.0%-100%), high-grade serous (99.1%) or endometrioid (97.8%) ovarian cancer, and urothelial carcinoma (92.6%-100%). A low CK19 positivity rate (0.1-10%) was seen in 5 of 122 tumor entities including hepatocellular carcinoma and seminoma. A comparison of tumor versus normal tissue findings demonstrated that upregulation and downregulation of CK19 can occur in cancer and that both alterations can be linked to unfavorable phenotypes. CK19 downregulation was linked to high grade (p=0.0017) and loss of estrogen receptor- and progesterone receptor-expression (p<0.0001 each) in invasive breast carcinoma of no special type. CK19 upregulation was linked to nodal metastases in neuroendocrine tumors and papillary thyroid carcinomas (p<0.05 each) and to poor grade in clear cell renal cell carcinoma (p<0.05). CK19 upregulation was particularly common in squamous cell carcinomas. We concluded that CK19 IHC might separate primary liver cell carcinoma from liver metastases, seminoma from other testicular tumors, and helps in the detection of early neoplastic transformation in squamous epithelium.

Prognostic significance of hormone receptor (ER/PR) status in endometrial carcinoma in black women: Implications with lymph node metastasis.

e18099 Background: Histological examination of tumor tissues is necessary for deciding if endometrial surgery should include lymph node sampling. Studies have demonstrated that loss of estrogen and progesterone receptors (ER/ PR) independently predicts lymph node metastasis and poor prognosis. African American females in comparison to their Caucasian counterparts are diagnosed at a later stage, with higher grade, poorer prognostic histological types and worse outcomes. Our study aimed to investigate if the correlation between hormone receptor status and lymph node metastasis holds true in African American females which could aid in treatment strategies. Methods: We reviewed African American females (n = 70) diagnosed with Endometrial Cancer at our institution from 2015-2018 including endometrioid/non-endometrioid histology, FIGO stage I-IV, grade 1-3 for ER/PR receptors and presence or absence of pelvic lymph nodes based on preoperative endometrial curettage and post-surgical pathology specimens. Statistical analyses were performed using Fisher’s Exact test exploring associations between categorical variables (ER/PR and Lymph nodes Metastasis). All statistical tests were two-sided and considered significant if P value &lt; 0.05. Results: The mean age of our sample was 61 ± 13 years. Mean BMI was 36±10kg/m2. Majority of our patients were postmenopausal (59 out of 70, 84%). FIGO Stage 1 and 2 constituted 72% (50 out of 70). Endometrioid histology constituted 39 out of 70 (55%) cases and non-endometrioid histology which includes serous, mixed and carcinosarcomas constituted 45% of the cases. Loss of estrogen receptor(ER-) was observed in 19 of 70 patients (27%) and loss of progesterone receptor(PR-) was seen in 22 patients (31%) which was significantly associated with lymph node metastasis (p value &lt; 0.05).On subgroup analyses significant association was observed between PR+ and absence of lymph node metastasis in cases of endometrioid cancer (p value &lt; 0.05) . Nevertheless, there was no statistical significant association (P value .10) between ER receptors and lymph node metastasis in patients with endometrioid cancer. In non-endometrioid histology both ER- and PR- were found to have statistical significant lymph node involvement. Conclusions: Loss of ER and PR receptors independently predicts lymph node metastasis in endometrial cancers in African American females.

Loss of Estrogen Receptors is Associated with Increased Tumor Aggression in Laryngeal Squamous Cell Carcinoma

Laryngeal squamous cell carcinoma (LSCC) responds to 17β-estradiol via estrogen-receptor (ER, transcribed from ESR1) dependent mechanisms, but is not recognized as a hormonally responsive cancer. 17β-estradiol production by LSCC cell lines UM-SCC-11A and UM-SCC-12 was examined. Wild type (WT) and ESR1-silenced LSCC cultures and xenografts were examined for 17β-estradiol responsiveness in vivo. 14 LSCC and surrounding epithelial samples at various pathological stages were obtained from patients; ERα and ERβ expression were verified using data from the total cancer genome atlas. UM-SCC-11A and UM-SCC-12 both produce 17β-estradiol, but only UM-SCC-12, not UM-SCC-11A, xenograft tumors grow larger in vivo in response to systemic 17β-estradiol treatments. ERα66 and ERα36 expression inversely correlated with clinical cancer stage and tumor burden. LSCC ERα66 expression was higher compared to surrounding epithelia in indolent samples but lower in aggressive LSCC. ERβ expression was highly variable. High ESR1 expression correlated with improved survival in LSCC. Loss of ERα66 expression inversely correlated with prognosis in LSCC. ERα66 may be a histopathological marker of aggression in LSCC.

AB066. 208. Loss of estrogen receptor (ER) through DNA methylation and alterations in molecular heterogeneity during endocrine treatment in ER-positive breast cancer

AB066. 208. Loss of estrogen receptor (ER) through DNA methylation and alterations in molecular heterogeneity during endocrine treatment in ER-positive breast cancer

Increase in hyaluronic acid degradation decreases the expression of estrogen receptor alpha in MCF7 breast cancer cell line.

The loss of estrogen receptor α (ERα) expression in breast cancer constitutes a major hallmark of tumor progression to metastasis and is generally correlated to a strong increase in Hyaluronic Acid (HA) turnover. The aim of our study was to search for a putative link between these two major events of breast cancer progression in the estrogen receptor-positive (ER+) MCF7 breast cancer cell line. The increase in HA turnover was performed by stable overexpression of the standard CD44 (CD44S) isoform and also by treatment with exogenous Hyaluronidase (Hyal) to allow an increase in HA catabolism. Stable overexpression of CD44S in MCF7 cells was correlated to a decrease in ESR1 gene expression, which did not lead to alteration of estrogen response. Moreover, our results showed that the exposure to exogenous Hyal stimulates the proliferation and strongly decreases the expression of ERα whatever the expression level of CD44 in the MCF7 cell line. The culture in the presence of Hyal led to the decrease in estrogens responsiveness and to hormonal therapy resistance. The effect on growth is correlated to the activation of MAPK/ERK and PI3K/Akt signaling pathways while the Hyal-induced down-regulation of ESR1 gene expression involves the activation of PI3K/Akt and NF-κB signaling pathways. Many of our data suggest that the effects of Hyal described here could be related to the activation of TLR signaling. Taken together, our results demonstrate that the increase in HA degradation could be involved in breast cancer progression and in resistance to hormonal therapy.

Breast cancer subtype discordance: impact on post-recurrence survival and potential treatment options

BackgroundRecent studies have shown that breast cancer subtype can change from the primary tumour to the recurrence. Discordance between primary and recurrent breast cancer has implications for further treatment and ultimately prognosis. The aim of the study was to determine the rate of change between primary and recurrence of breast cancer and to assess the impact of these changes on survival and potential treatment options.MethodsPatient demographics were collected on those who underwent surgery for breast cancer between 2001 and 2014 and had a recurrence with biopsy results and pathology scoring of both the primary and recurrence.ResultsOne hundred thirty two consecutive patients were included. There were 31 (23.5%) changes in subtype. Discordance occurred most frequently in luminal A breast cancer (n = 20), followed by triple negative (n = 4), luminal B (n = 3) and HER2 (n = 3). Patients who changed from luminal A to triple negative (n = 18) had a significantly worse post-recurrence survival (p < 0.05) with overall survival approaching significance (p = 0.064) compared to concordant luminal A cases (n = 46). Overall receptor discordance rates were: estrogen receptor 20.4% (n = 27), progesterone receptor 37.7% (n = 50) and HER2 3% (n = 4). Loss of estrogen receptor and progesterone receptor was more common than gain (21 vs. 6 (p = 0.04) and 44 vs. 6 (p = 0.01) respectively). Nine patients (6.8%) gained receptor status potentially impacting treatment options.ConclusionDiscordance in subtype and receptor status occurs between primary and recurrent breast cancer, ultimately affecting survival and potentially impacting treatment options.

Impact of oestrogen and progesterone receptor expression in the cancer cells and myometrium on survival of patients with endometrial cancer

Loss of oestrogen receptor (ER) and progesterone receptor (PR) expression in endometrial cancer cells indicates poor prognosis. The aim of this study was to determine the correlation of ER and PR expression in cancer cells and the surrounding myometrium with the disease progression. Immunohistochemical expression of ER and PR was detected in cancer and myometrial cells of patients with EC. ER was detected in 65.2% of cancer cells and in 88.4% of myometrial cells. PR was detected in 59.4% of cancer cells and in 84.1% of myometrial cells. The 5-year overall survival (OS) was 76.8%. Patients with ER and PR negative EC had a shorter period until recurrence (p = .013 and .043) and shorter OS (p = .011 and .066) than those with ER and PR positive cancer. Negative ER and PR status in EC has an impact on recurrence and poor OS. The status of hormone receptors in myometrium may be useful in disease prognosis.Impact StatementThe status of hormone receptors in endometrial cancer has been the subject of numerous studies and loss of hormone receptors indicates higher tumor grade and higher clinical stage, lympho-vascular space invasion and deeper myometrial invasion. Although, the communication between the endometrium and myometrium is crucial under physiological conditions, the status of hormone receptors in the myometrium and its significance in cancer progression is poorly studied.Our results showed that loss of ER in the myometrium indicate poor prognosis. The assessment of hormone receptor status in myometrium might be useful in predicting the course of the disease. Results of our research support the theory that stromal and myometrial cells may contribute to tumorigenesis in endometrial cancer.Better understanding of ER/PR expression in myometrial cells is needed, and our research opens new possibilities for identification of key pathways and new potential target molecules in EC prognosis and treatment. It is probable that future classification of endometrial cancer will rely on molecular sub-typing, where the status of hormone receptors in the myometrium might play an important role.

Oestrogen receptor negativity in breast cancer: a cause or consequence?

Endocrine resistance, which occurs either by de novo or acquired route, is posing a major challenge in treating hormone-dependent breast cancers by endocrine therapies. The loss of oestrogen receptor α (ERα) expression is the vital cause of establishing endocrine resistance in this subtype. Understanding the mechanisms that determine the causes of this phenomenon are therefore essential to reduce the disease efficacy. But how we negate oestrogen receptor (ER) negativity and endocrine resistance in breast cancer is questionable. To answer that, two important approaches are considered: (1) understanding the cellular origin of heterogeneity and ER negativity in breast cancers and (2) characterization of molecular regulators of endocrine resistance. Breast tumours are heterogeneous in nature, having distinct molecular, cellular, histological and clinical behaviour. Recent advancements in perception of the heterogeneity of breast cancer revealed that the origin of a particular mammary tumour phenotype depends on the interactions between the cell of origin and driver genetic hits. On the other hand, histone deacetylases (HDACs), DNA methyltransferases (DNMTs), miRNAs and ubiquitin ligases emerged as vital molecular regulators of ER negativity in breast cancers. Restoring response to endocrine therapy through re-expression of ERα by modulating the expression of these molecular regulators is therefore considered as a relevant concept that can be implemented in treating ER-negative breast cancers. In this review, we will thoroughly discuss the underlying mechanisms for the loss of ERα expression and provide the future prospects for implementing the strategies to negate ER negativity in breast cancers.

Abstract 4396: Circulating CAFs and CAF-secreted factors may be indicative of breast cancer metastasis

Abstract Tumor metastasis is the main cause of breast cancer mortality. Increasing evidence demonstrates stromal cells play pivotal roles in promoting breast cancer progression and metastasis. Breast cancer stroma is comprised mainly of Cancer Associated Fibroblasts (CAFs). CAFs secrete various growth factors and cytokines that promote breast cancer progression and metastasis; one of these factors is SDF-1 (CXCL12), a soluble chemokine that is promotes chemotaxis and motility, and facilitates cancer cell motility and angiogenesis. CAFs also secrete soluble factors that activate ERK 1/2 MAPK signaling in breast cancer cells, which has been shown to promote loss of estrogen receptor (ER) in luminal breast cancer cells. Hyperactivation of MAPK signaling (hMAPK) also associates with aggressive, basal-like and HER2-positive breast cancer and poor prognosis. Recently, we identified a patient-derived hMAPK-microRNA signature indicative of poor clinical outcome that contains microRNAs known to regulate breast cancer associated genes. The vast majority of ER- breast cancers display this microRNA signature, as do a subset of ER+ breast cancers with poorer clinical outcome. We also discovered that the breast cancers that exhibit this microRNA signature display high stromal and immune infiltrate scores, suggesting that breast cancer stroma provides important contributions to this microRNA signature and the poor clinical outcomes associated with it. To study the role of CAFs and CAF-secreted factors in breast cancer progression and metastasis, we established primary breast CAF lines from ‘indolent’ breast cancers (Luminal A), and from ‘aggressive’ breast cancers (ER-/HER2 amplified; triple negative). We have demonstrated that these CAFs differentially express several members of the hMAPK-microRNA signature compared to cultured primary breast cancer cells, supporting the contribution of stroma to the signature. Importantly hMAPK-microRNAs secreted from “aggressive” CAFs can be taken up by breast cancer cells, whereupon they repress their targets. Normal human mammary fibroblasts (HMFs) and ‘indolent’ CAFs do not secrete these microRNAs. We identified a novel class of circulating cells in the blood of breast cancer patients with metastases -CAFs (cCAFs). Patients without metastases did not have these cCAFs - suggesting cCAFs may be “aggressive” CAFs that facilitate breast cancer metastasis. Patients with overt metastasis and elevated counts of cCAFs have significantly higher levels of circulating SDF-1 in their plasma, as well as differential circulating microRNAs, and specifically hMAPK-microRNAs also found secreted by “aggressive” CAFs. Our results suggest there is a hierarchy of CAFs whereby “aggressive” tumors establish “aggressive” CAFs to facilitate metastasis. We also establish a clear link between circulating CAFs and CAF-secreted factors such as SDF-1 and microRNAs with breast cancer metastasis. Citation Format: Sanket H. Shah, Phil Miller, Leah Machlin, Kelsie Medina-Saenz, Ritesh Parajuli, Marc E. Lippman, Dorraya El-Ashry. Circulating CAFs and CAF-secreted factors may be indicative of breast cancer metastasis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4396.

Abstract A20: Decreased nuclear expression of the FRY protein identifies a subset of breast cancers with poor clinical outcomes

Abstract The goal of the present study was to determine if decreased nuclear expression of the human FRY protein can serve as a biomaker of breast cancer progression and outcomes. We previously identified the rat Fry gene as a putative mammary carcinoma susceptibility (Mcs) gene. We further demonstrated that FRY expression was decreased in several human breast cancer cell lines. Moreover, ectopic expression of wildtype Fry suppressed tumorigenicity of the triple negative MDA-MD-231 breast cancer cells in vitro and in vivo by promoting epithelial cell differentiation. To evaluate the contribution of altered FRY expression to the clinical progression of human breast cancer, we first compared FRY mRNA expression in &amp;gt;4,800 clinically annotated human breast cancers using DNA microarray data from 19 distinct cohorts available from the Oncomine 3.0 Cancer Profiling Database. The analysis indicated that decreased FRY mRNA was significantly correlated with poorly differentiated tumor histopathology, increased Elston tumor grade, and triple negative status (loss of estrogen, progesterone and Her2 receptors). Using commercially available breast cancer tissue microarrays, we used semi-quantitative immunohistochemistry and quantitative image analysis to compare FRY protein expression in normal mammary tissue and tumors. The results indicated that loss of FRY expression was a frequent event during progression of mammary carcinomas. We therefore analyzed &amp;gt;1200 clinically annotated breast cancers represented on tissue microarrays from commercial suppliers and the National Cancer Institute (NCI) Cancer Diagnosis Program, including the Breast Cancer Diagnosis and Progression panels. FRY protein expression was evaluated by immunohistochemical staining and was graded on a scale of 0 to 4 (0, +, ++, +++, ++++). The analysis indicated that decreased nuclear expression of the FRY protein was significantly correlated with poorly differentiated histopathologies, increasing Elston grade and loss of the estrogen receptor. Tumors with decreased FRY expression were also more likely to metastasize and were associated with decreased tumor-free survival. These findings indicated that loss of nuclear FRY expression identifies a subset of breast cancer with poor clinical outcomes, suggesting that FRY may serve as an early biomarker for breast cancer progression and prognosis. Citation Format: Helmut Zarbl, Jessica C. Graham, Norio Takizawa, Mingzhu Fang, Zhihong Gong, Brian Estrella, Xuefeng Ren. Decreased nuclear expression of the FRY protein identifies a subset of breast cancers with poor clinical outcomes. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr A20.

S100A8/A9 is associated with estrogen receptor loss in breast cancer.

S100A8 and S100A9 are calcium-binding proteins that are secreted primarily by granulocytes and monocytes, and are upregulated during the inflammatory response. S100A8 and S100A9 have been identified to be expressed by epithelial cells involved in malignancy. In the present study, the transcriptional levels of S100A8 and S100A9 were investigated in various subtypes of breast cancer (BC), and the correlation with estrogen receptor 1 (ESR1) and GATA binding protein 3 (GATA3) gene expression was evaluated using microarray datasets. The expression of S100A8 and S100A9 in BC cells was assessed by reverse transcription-polymerase chain reaction (RT-PCR). The regulation of ESR1 and GATA3 by administration of recombinant S100A8/A9 was examined in the BC MCF-7 cell line using quantitative (q)PCR. The association between S100A8 and S100A9 and overall survival (OS) was investigated in GeneChip® data of BC. The expression levels of S100A8 and S100A9 were higher in human epidermal growth factor receptor 2 (Her2)-amplified and basal-like BC. The messenger (m)RNA levels of S100A8 and S100A9 were inversely correlated with ESR1 and GATA3 expression. S100A8/A9 induced a 10-fold decrease in the mRNA levels of ESR1 in MCF-7 cells. Poor OS was associated with high expression levels of S100A9, but not with high expression levels of S100A8 in BC. In conclusion, strong expression and secretion of S100A8/A9 may be associated with the loss of estrogen receptor in BC, and may be involved in the poor prognosis of Her2+/basal-like subtypes of BC.

Undifferentiated Carcinoma of the Endometrium: An Expanded Immunohistochemical Analysis Including PAX-8 and Basal-Like Carcinoma Surrogate Markers.

Undifferentiated carcinoma of the endometrium (UCAe) is an aggressive, underrecognized high-grade carcinoma that can occur either in pure form or in conjunction with low-grade endometrioid adenocarcinoma (i.e. dedifferentiated carcinoma). The typical solid growth pattern of UCAe can create a diagnostic dilemma as it is frequently misinterpreted as the solid component of an endometrial carcinoma or as a sarcoma. In addition, the high nuclear:cytoplasmic ratio, high mitotic index, and geographic necrosis are reminiscent of basal-like carcinoma of breast (BLCB). This study was undertaken to determine the role of a selected group of immunomarkers in the distinction of UCAe from other endometrial carcinomas, and assess the expression of DNA mismatch repair proteins, and surrogate BLCB immunomarkers in this type of tumor. Cases of UCAe were stained with antibodies against keratin cocktail, CK8/18, PAX-8, and estrogen receptor: 35 cases; progesterone receptor and Her-2/neu: 33 cases; CD44, e-cadherin, p16, and p53: 32 cases; and CK5/6, EGFR, and c-Kit: 18 cases. In addition, mismatch repair protein markers MLH1, MSH2, MSH6, and PMS2 were performed in 34 cases. We found that PAX-8 expression was lost in most cases (83%). In addition, estrogen and progesterone receptors were negative in 83% and 82% of cases, respectively. Seventy-seven percent of cases were positive for keratin cocktail and keratin 8/18, whereas only 11% of cases were positive for keratin 5/6. p16 was diffusely positive in 34% of cases, whereas p53 was expressed in >75% of the tumor cells in 31% of cases. MLH1 and PMS2 were concurrently lost in 50% of cases, whereas MSH2 and MSH6 were lost in 1 case (3%). E-cadherin and CD44 were completely lost in 50% of cases, whereas Her-2/neu was negative in all cases. EGFR was negative in 67% of cases, whereas 22% of cases showed diffuse membranous staining for this marker. UCAe is a high-grade carcinoma of Müllerian origin which tends to be negative for PAX-8. The loss of this marker appears to be a more reliable discriminator than the loss of keratin expression in the differential diagnosis with endometrioid carcinoma or serous carcinoma. UCAe tends to be diffusely positive for p53, but patchy positive for p16. Although UCAe appears to share not only some histologic features with BLCB, but also some of its immunohistochemical features (loss of estrogen receptor, progesterone receptor, and Her-2/neu, a tendency to loose e-cadherin and to express CD44), UCAe appears not to be related to BLCB because it usually lacks the expression EGFR, CK5/6, and c-Kit.