Estrogen Receptor Positive Tumors Research Articles

Abstract PO3-01-14: Impact of detection mode, tumor characteristics and additional treatment on adherence to adjuvant endocrine therapy after breast cancer in Sweden

Abstract Background: Adjuvant endocrine therapy (AET) reduces recurrence and mortality in women with estrogen receptor (ER) positive early breast cancer (EBC). Adherence to AET has been shown to be lower than expected with risk of worse longtime prognosis. In a nationwide study in Sweden women with breast cancer diagnosis 2008-2010 adherence to AET at 5-year follow-up was estimated at 82,5% which is considered good related to recent published studies (manuscript is under review). Differences between regions and age-groups were shown. Present study investigates the impact on adherence of detection mode, tumor characteristics and additional treatment. Methods: Through the Swedish Cancer Registry (SCR) women with a first primary EBC diagnosed 2008-2010 were identified. From the Swedish National Breast Cancer Registry (NKBC) individual tumor and treatment data were collected. Included in the study was patients with ER positive tumors > 10 mm without distant metastasis at diagnosis. Through the Swedish Prescription Registry dispensed treatment from pharmacies was extracted and medication possession rate (MPR) was calculated as number of dispensed doses divided by treatment duration in days. Good adherence to treatment in a patient was set at MPR ≥80%. Adherence at 3- and 5-year follow-up by detection mode, tumor characteristics and additional treatment was estimated. Results: Out of 21 016 women with a first primary BC 2008-2010 registered in the NKBC 10 176 met the inclusion criteria for the study. Among screen-detected tumors the 3- and 5-year adherence were 90,2 respectively 85,1% and 85,4 respectively 78,4% in the non-screen-detected group. In the screen-detected group the tumors were smaller and more often node negative. There was a significantly increased 5-year adherence in patients with stage III disease compared to those with stage I disease (84,1 vs 81,3 %) but not significant at 3-year (89,2 vs 87,3). Adjuvant chemotherapy significantly increased 3- (91.0% vs 85.8%) and 5-year (85.7% vs 79.3%) adherence among screen-detected and non-screen-detected respectively. Conclusions: Adherence to AET in Sweden was good, although there were differences depending on detection mode, tumor stage and additional adjuvant treatment. Stage and additional medical adjuvant treatment depend on each other and should not be seen as independent factors for adherence for AET. Citation Format: Anne Andersson, Anna Von Wachenfeldt, Lennarth Nyström, Frida Isaksson, Pihla Ruohonen. Impact of detection mode, tumor characteristics and additional treatment on adherence to adjuvant endocrine therapy after breast cancer in Sweden [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-01-14.

Abstract PO4-27-12: Increasing DAXX as a Novel Approach to Inhibit Breast Cancer Stem Cells and Estrogen Receptor-positive Tumor Recurrence

Abstract Background: Resistance to endocrine therapy (ET; tamoxifen, aromatase inhibitors, AI, or fulvestrant) in ER+ breast cancer (BC) could be due to survival of breast cancer stem cells (BCSCs). BCSCs contribute to relapse of ER+ BC. We discovered a novel and potent anti-BCSC gene, Death Associated Protein 6 (DAXX) through a pre-surgical biomarker window study combining ET plus a Notch inhibitor [MK-0752, a g-secretase inhibitor (GSI)]. We also found that ET enhances the BCSC population by decreasing DAXX protein levels. In this current study, we measured DAXX protein levels in ER+ PDX tumors and human tumor tissues treated with ET. We determined the mechanism by which ET regulates DAXX expression at the level of protein stability. The goal of this study was to identify novel therapeutic strategies to prevent ET-mediated degradation of the DAXX protein, eliminate BCSCs, and prevent ER+ recurrence. Methods: ER+ BC cell lines (MCF-7 and T47D), an ER+ PDX BCM 5097 tumor line, and human breast cancer tumor samples were used. The ER+ PDX tumor (BCM 5097) was implanted into NSG mice and passaged into female athymic, nude mice-supplemented with an estradiol capsule. After one week, the estradiol capsule was removed from 10 mice, to mimic the use of AI. The remaining 10 mice retained the estradiol capsule. Tumor area was measured weekly up to 50 weeks. Western blotting detected ERa, the stem cell factor Notch4, and DAXX levels. Immunohistochemistry (IHC) detected DAXX protein levels. ER+ human tumor samples were collected prior to and after neoadjuvant treatment with ET (N=11) or chemotherapy (N=14) and stained for the DAXX protein. Correlations were made between DAXX and Ki67. Full length and deletion mutants of DAXX were expressed in ER+ BC cells. BCSCs were measured using the mammosphere forming assay. Liquid chromatography followed by mass spectrometry were used to detect phosphorylated residues of DAXX in response to ET. Kinases that phosphorylate these residues on DAXX were identified using PhosphoSite.org. ER+ cells were treated with kinase inhibitors for AURKA (alisertib), AURKB (barasertib), CK1 (CK-IN-1), or CK2 (CX-4945) and DAXX protein was detected by western blotting. BCSC survival was measured using mammosphere forming assay. Based on results from the kinase inhibitor screen, barasertib was selected for pre-clinical testing in mice. ER+ MCF-7 xenograft tumors were treated with vehicle or barasertib in vivo up to 120 days. Results: DAXX protein levels decreased in ER+ PDX tumors in response to ET. Similar trends were detected in human tumor tissue after ET or chemotherapy alone. DAXX protein levels inversely correlated with Ki67 in ER+ human tumor samples. Analysis of DAXX deletion mutants demonstrated that amino acids 400-740 of the DAXX protein were required to inhibit BCSC survival. This region of the DAXX protein had high levels of phosphorylated residues in response to ET. Only the AURKB (barasertib) inhibitor increased DAXX protein expression and inhibited BCSCs in a DAXX-dependent manner. Barasertib inhibited growth of ER+ MCF-7 tumor xenografts compared to the vehicle control. After treatment ceased, tumor recurrence was measured up to 120 days. Regrowth of tumors was partially delayed in response to barasertib. Conclusions: ET decreased DAXX protein levels in ER+ PDX and human tumors. Downregulation of the DAXX protein by ET was through activation of AURKB and hyper-phosphorylation of DAXX which resulted in protein degradation and enhanced survival of BCSCs. Therefore, Inhibition of AURKB using barasertib partially restored DAXX expression, inhibited BCSCs, and delayed tumor recurrence. A combination approach of ET plus an AURKB inhibitor might be a novel therapeutic strategy to prevent ER+ breast cancer relapse by increasing DAXX in order to eradicate BCSCs. Support: Breast Cancer Research Foundation Citation Format: Clodia Osipo, Debra Wyatt, Kathy Albain. Increasing DAXX as a Novel Approach to Inhibit Breast Cancer Stem Cells and Estrogen Receptor-positive Tumor Recurrence [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-27-12.

Prognostic value of molecular classification in stage IV endometrial cancer

ObjectivesMultiple studies have proven the prognostic value of molecular classification for stage I–III endometrial cancer patients. However, studies on the relevance of molecular classification for stage IV endometrial cancer patients...

Male Breast Cancer: An Updated Review of Patient Characteristics, Genetics, and Outcome.

Male breast cancer (MBC) is a rare entity, underrepresented in population studies and clinical trials, resulting in management of MBC to be informed by current research on female breast cancer (FBC). A literature review was conducted by accessing relevant articles on 2 databases, by searching keywords "male breast cancer". A total of 29 articles from year 2011 to 2022 were selected for this review. The authors found that male breast cancer generally occurs later in life with higher stage, higher grade, and more estrogen receptor (ER) positive tumours. Most of the studies noted the mean age for MBCs at the time of presentation as >60 years. Risk factors for male breast cancer include family history, obesity, lower physical activity, and syndromes like the Klinefelter syndrome. Positive family history is much higher in MBC compared to FBC (30.9 vs. 18.4%). BRCA 2 cancers constitute a higher proportion compared to FBCs. A lot of genetic mutations have been observed. Some show promise to assess disease-specific survival and proliferative rate like TWIST1 and RUNX3, among others. MBCs usually present with a palpable lump in central region, with a bigger size and chance of nodal involvement and metastasis compared to FBCs. They are mostly infiltrating ductal type and hormone receptor positive, with worse histological grade. Treatment usually follows the same principles as FBCs (systemic therapy, surgical excision, and radiotherapy), with poorer prognosis to same treatment approach, possibly owing to its advanced stage at presentation. This is a rare entity which requires further research to ascertain need for different management approach than FBCs.

Abstract 1201: Pervasive HLA disruption fuels cancer evolution

Abstract Disruption of the human leukocyte antigen (HLA) molecules has important implications for immune evasion and tumor evolution. However, although genomic loss of HLA is frequent in non-small cell lung cancer (NSCLC) and in some types of breast cancer, the extent and importance of transcriptomic disruption to HLA presentation, including transcript repression and alternative splicing (AS), remains unclear. To address this, we developed MHC Hammer, a tool to call HLA allele-specific mutations, loss of heterozygosity (LOH), transcriptional repression, and alternative splicing. We first assessed HLA allelic expression and the prevalence of AS in normal tissue. Applying MHC Hammer to 510 normal lung samples and 407 normal breast samples from the Genotype-Tissue Expression cohort, we identified extensive heterogeneity in HLA allelic expression depending on both the allele and tissue type. In addition, HLA allelic AS, occurring in at least 20% of the allele transcripts, was observed in 19% of normal lung and 13% of normal breast samples. Next, we applied MHC Hammer to 413 TRACERx NSCLCs and 743 TCGA breast tumours. While mutations were uncommon, HLA LOH was a frequent event, occurring in 32% of lung adenocarcinomas (LUAD) and 57% of lung squamous cell carcinomas (LUSC). We observed a similar rate of HLA LOH in estrogen receptor positive (ER+) and negative (ER-) tumors (ER+: 14%, ER-: 15%), while in triple negative breast tumours we observed a higher rate of LOH (31%). Due to the heterogeneity in HLA expression observed in the normal samples, to measure tumor specific HLA repression and alternative splicing, we restricted our analysis to tumours with WES, tumor and tumor-adjacent normal RNAseq data. This resulted in a cohort of 88 NSCLCs from the TRACERx study as well as 44 ER+ breast tumours from the TCGA cohort. 61% of LUAD, 75% of LUSC and 35% of ER+ tumours harbored class I HLA transcriptional repression that could not be explained by underlying genomic loss. 27% of LUAD, 7% of LUSC and 55% of ER+ breast tumor patients exhibited no HLA disruption. Tumor specific AS affecting HLA exons 2, 3 or 4, potentially disrupting HLA antigen presentation, was identified in 8% of LUADs, 11% of LUSC and 7% of ER+ breast tumours. Tumor specific AS of exon 5, yielding soluble HLA molecules without a transmembrane domain was identified in 20% of LUAD, 14% of LUSCs and 18% of ER+ breast tumours. LUAD and LUSC tumor regions without HLA LOH or repression were enriched for somatic HLA AS events and alleles with somatic HLA AS had a higher neoantigen burden than those without. We found no relationship between survival and HLA expression or AS in the normal tissue. However, consistent with the importance of HLA dysfunction in tumor evolution, LUADs with low HLA expression in at least one tumor region were associated with shorter disease-free survival, HLA LOH was more common in LUAD’s that metastasized and HLA repression was enriched in LUSC primary regions seeding metastases. Citation Format: Clare Puttick, Thomas P. Jones, Michelle Leung, Oriol Pich, Felipe Galvez Cancino, Jiali Liu, Manuel Varas-Godoy, Andrew Rowan, Carlos Martinez-Ruiz, Robert Bentham, Krijn K. Dijkstra, Rachel Rosenthal, Nnennaya Kanu, Kevin Litchfield, Roberto Salgado, David A. Moore, Peter Van Loo, Mariam Jamal-Hanjani, Sergio A. Quezada, Nicholas McGranahan, Charles Swanton. Pervasive HLA disruption fuels cancer evolution [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1201.

Abstract 434: Understanding metabolic heterogeneity of estrogen receptor positive metastatic breast tumors using spatial transcriptomics and geographic information systems approaches

Abstract The tumor microenvironment consists of numerous cell populations showing diverse metabolic preferences, causing metabolic heterogeneity and therapy resistance. Fulvestrant, a clinically approved selective estrogen receptor (ER) degrader, is the standard of care for ER-positive breast cancer, yet resistance has been reported among metastatic breast cancer patients. In our previous studies, we showed such resistance is attributable to metabolic heterogeneity in the tumor microenvironment. Despite metabolic heterogeneity posing a common challenge in cancer research, current methods, such as histology and Magnetic Resonance Imaging, cannot explore nutrient uptake at transcript-level resolution with spatial context, making it difficult to understand the heterogeneity that affects cellular growth and metabolism. Visium Spatial Transcriptomics, a method that shows regional gene expression, is emerging as a powerful spatial analysis tool, and we integrated this method with geographic information systems to investigate the tumor microenvironment heterogeneity and highlight the importance of cell-microenvironment interactions. We developed a computational pipeline to visualize hotspots of spatially relevant genes using breast cancer metastatic samples from xenograft models and human patients. This pipeline integrates Visium tools and geospatial methods for processing transcriptome profiling data. We analyzed the degree of local spatial association based on Local Moran’s I to estimate regions with significant co-localization of single and multiple gene expressions within a tumor. We then leveraged the co-localized areas in the process of biomarker identification and revealing biological pathways. We also used unsupervised clustering of the complete transcriptome profiles to test the resemblance between identified clusters and hotspots as well as any disruption in the spatial structure of clusters in response to therapy or metastasis. Gene set enrichment analysis of these clusters revealed several glucose or lipid metabolism pathways, and mapping of the genes corresponding to each pathway showed mutually exclusive gene expression patterns in the sample. We also observed minimal co-localization of two metabolic pathways but extensive co-localization between metabolic genes and ER target genes, suggesting distinct metabolic preferences and spatial distributions of endocrine therapy resistant cell populations. Altogether, our analysis pipeline established a computational framework for characterizing the link between tumor microenvironment, local metabolism, and changes in transcriptional response to external stimuli. Our findings support the development of strategies to target different endocrine resistant regions using a combination of ER and metabolic signaling inhibitors. Citation Format: Jin Young Yoo, Sabrina Akter, Nandana Varma, Atharva Jain, Aiman Soliman, Zeynep Madak Erdogan. Understanding metabolic heterogeneity of estrogen receptor positive metastatic breast tumors using spatial transcriptomics and geographic information systems approaches [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 434.

Abstract 4342: Breast cancer multi-omic single cell profiling identifies key progressive disease markers

Abstract Breast cancer is one of the most diagnosed cancers and has a complex tumor microenvironment (TME). Breast tumors are subtyped by receptor expression, and all have a high degree of cellular heterogeneity. We analyzed tumors from 8 individuals with early-stage breast cancer. The patients had either triple negative breast (TNBC) or an estrogen receptor (ER) positive cancer. The tumors and normal adjacent tissue were profiled by single-nuclei gene expression (GEX) and ATAC data using the 10X multi-ome methodology. Gene set analysis identified specific epithelial clusters associated with the estrogen early/late response, epithelial mesenchymal transition, and several key cytokine pathways. From those epithelial cells, a subset highly proliferative in nature emerged. It consists of both ER positive and TNBC tumor types enriched for specific DNA transcriptional motifs including those for stem cell (Yamanaka) factors and other cellular proliferation motifs. GEX proliferation markers such as MKi-67 were also highly expressed. Overlaying GEX and ATAC data, it was possible to filter the numerous differentially expressed genes and peaks into a subset of genes specific to progressive breast cancer within the proliferative cluster. Those genes included ESRP1, POP4, CCNE1, and AAMDC all of which are associated with progressive breast cancer. To further elucidate the complexity of the TME, we examined clusters of fibroblasts from our dataset. A difference in cancer associated fibroblast marker gene expression was related to the level of ER expression expressed in the tumor epithelium. In general, ER-low tumor fibroblasts had higher expression of immune cancer associated fibroblasts (iCAFs) markers while ER-positive and TNBC tumors had greater expression of extracellular matrix associated markers (emCAFs). Specific clusters were enriched for emCAFs and iCAFs representing another critical part of the TME. In conclusion, using a multi-modal single-cell approach to the TME allowed the identification of an aggressive cluster of epithelial cells marked by genes for cellular proliferation and noted therapeutic targets while also characterizing other critical TME components especially, CAFs. Citation Format: Hugh Russell, Manisha Rao, Grant Duclos, Rogelio Aguilar, Nick Barkas, Megan Callahan, Ojasvi Chaudhary, Peter Gathungu, Chris Rands, Varsha Shankarappa, Daniel Stetson, Asaf Rotem, Maurizio Scaltriti, Brian Dougherty. Breast cancer multi-omic single cell profiling identifies key progressive disease markers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4342.

Abstract 5534: Needle biopsy induces pro-metastatic progression and systemic dissemination of estrogen receptor-positive breast cancer cells through the COX-2/PGE2 cascade

Abstract Breast cancer is the most commonly diagnosed malignancy in the U.S. and over two-thirds of invasive breast cancer cases are diagnosed early stage. However, 15-year survival rates for early-stage breast cancer remain in the range of 70-77%. Mounting evidence showed an increased risk of disease progression and mortality among early-stage breast cancer patients whose surgery was delayed over 60 days after the diagnostic biopsy. Yet, the mechanism(s) underlying the rapidly increased mortality due to delay of surgery after diagnosis remains unknown. Our cohort analyses of early-stage breast cancer patients reveal the emergence of a significantly rising mortality risk when the biopsy-to-surgery interval was extended beyond 53 days. Additionally, histology of post-biopsy tumors shows prolonged retention of a metastasis-permissive wound stroma dominated by M2-like macrophages capable of promoting cancer cell epithelial-to-mesenchymal transition and angiogenesis by secretion of transforming growth factor beta 1 (TGF-β1) and vascular endothelial growth factor (VEGF). We show that needle biopsy promotes systemic dissemination of cancer cells through a mechanism of sustained activation of the cyclooxygenase-2 (COX-2)/prostaglandin E2 (PGE2) cascade, which favors M2-polarization and its associated pro-metastatic changes, but are abrogated by oral treatment with COX-2 inhibitors in estrogen receptor-positive (ER+) syngeneic mouse tumor models. Therefore, we conclude that needle biopsy of ER+ breast cancer provokes progressive pro-metastatic changes, which may explain the mortality risk posed by surgery delay after diagnosis. Also, our data may provide a rationale for a pharmacologic strategy to inhibit the COX-2/PGE2 cascade in cases when delay of surgery is unavoidable. Citation Format: Hiroyasu Kameyama, Macall Leslie, Reese Simmons, Yunguang Sun, Misung Yi, Priya Dondapati, John F. Langenheim, Kar-Ming Fung, Inna Chervoneva, Hallgeir Rui, Takemi Tanaka. Needle biopsy induces pro-metastatic progression and systemic dissemination of estrogen receptor-positive breast cancer cells through the COX-2/PGE2 cascade [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5534.

Abstract B031: High risk no specific molecular profile (HR-NSMP) endometrial cancer can be stratified into three subgroups based on tumor grade and estrogen receptor status with differing clinicopathologic characteristics and outcomes

Abstract Introduction: Recent publications have identified ‘low risk’ (LR-NSMP) and ‘high risk’ no specific molecular profile (HR-NSMP) endometrial cancers (ECs). Although LR-NSMP (low grade (G1/2) estrogen receptor (ER) positive tumors have extremely low rates (<2%) of death from disease and are considered candidates for de-escalation of adjuvant therapy, little is known about the newly defined entity of HR-NSMP (G3 and/or ER-negative), including optimal management, with 27% of HR-NSMP patients dying from their disease We aimed to perform in-depth characterization of a cohort of HR-NSMP ECs to identify additional prognostic or predictive features that may inform management. Methods: Clinicopathologic data collection, immunohistochemistry (IHC), and next generation sequencing was performed on a cohort of 148 HR-NSMP ECs testing for associations with outcomes. Three subgroups of HR-NSMP were assessed: i) low grade without any ER expression (G1/2ER-, n=40), ii) high grade with ER (G3ER+, n=67), and iii) high grade without ER expression (G3ER-, n=41). Results: In a univariate analysis, advanced stage (III/IV), positive lymph node (LN) status, no LN assessment, and PIK3CA mutations were associated with inferior progression-free survival (PFS), disease specific survival (DSS), and overall survival (OS). HR-NSMP patients with positive LNs had a hazard ratio (HR) of death of 7.55 (95% CI: 3.41−16.73) compared to node negative. Additionally, patients with no LN assessment had an increased risk of death with a HR of 2.33 (95% CI: 1.12−4.85) compared to node negative. Within the three HR-NSMP subgroups, the worst OS (HR 7.06, CI 4.12-12.09, p< 0.001), DSS (HR 20.43, CI 10.14-41.19, p< 0.001), and PFS (HR 3.69, CI 1.59-8.63, p<0.001) were observed in patients with both adverse features (G3ER-). PIK3CA mutations were found in 32.7% of all HR-NSMP tumors tested (G1/2ER- 45.5%, G3ER+ 42.3%, G3ER- 6.7%) and associated with inferior OS (HR 4.84, CI 1.21-19.41, p=0.014), DSS (HR 4.84, CI 1.21-19.41, p=0.014), and PFS (HR 3.22, CI=1.08-9.66, p=0.028). There was a trend towards decreased OS and DSS in HR-NSMP patients with overexpression of HER2 IHC. L1CAM IHC overexpression and CTNNB1 mutation status were not associated with outcomes when assessed across all HR-NSMP or within subgroups. Conclusion: Among HR-NSMP ECs, the worst clinical outcomes were observed in patients harboring both high grade and ER negative tumors. In contrast to LR-NSMP ECs where patients who had no lymph node assessment had excellent outcomes mirroring node negative status, HR-NSMP patients who had no nodal assessment had a high rate of recurrence and death from disease suggesting the importance of comprehensive staging in this cohort. PIK3CA appears to be a prognostic stratification feature within HR-NSMP and represents a potential therapeutic pathway. More work is needed to understand the association of HER2 expression and ER status and whether HER2 -targeted therapies represent actionable opportunities for these patients. Citation Format: Andrea Neilson, Amy Jamieson, Derek Chiu, Samuel Leung, Amy Lum, Jennifer Pors, Stefan Kommoss, Aline Talhouk, David G. Huntsman, Naveena Singh, C. Blake Gilks, Jessica N. McAlpine. High risk no specific molecular profile (HR-NSMP) endometrial cancer can be stratified into three subgroups based on tumor grade and estrogen receptor status with differing clinicopathologic characteristics and outcomes [abstract]. In: Proceedings of the AACR Special Conference on Endometrial Cancer: Transforming Care through Science; 2023 Nov 16-18; Boston, Massachusetts. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(5_Suppl):Abstract nr B031.

Synergistic effect of Erythrina variegata L. and tamoxifen against the growth of breast cancer cells

Breast cancer is the most common malignancy and the second leading cause of death in women. While tamoxifen (TAM) remains to be a first-line agent for prevention and treatment of estrogen-receptor (ER) positive breast tumors, TAM resistance represents one of the biggest clinical challenges in breast cancer therapy. Therefore, there has been growing interest in the identification of novel agents including those of plant origin that enhance the TAM sensitivity of breast cancer cells. In the present study, we aimed to evaluate the synergistic effect of Erythrina variegata extract (EVE) and TAM on the suppression of breast cancer cell growth using MCF-7 cell model and conventional bioassays such as MTT and colony formation assay, cell cycle analysis, and immunocytochemistry. Interestingly, MTT assay in combination with Chou-Talalay analysis demonstrated the potent synergism between EVE and tamoxifen against MCF-7 cell survival. Additionally, co-treatment of MCF-7 cells with EVE and TAM more significantly suppressed colony formation, cell cycle progression, and the nuclear expression of proliferative marker Ki67 compared to TAM alone. These findings imply that EVE potentiates the anti-proliferative effect of TAM in MCF-7 breast cancer cells and may become a promising agent in the treatment of ER positive breast cancer.

Sex-specific outcomes in cancer therapy: the central role of hormones.

Sex hormones play a pivotal role in modulating various physiological processes, with emerging evidence underscoring their influence on cancer progression and treatment outcomes. This review delves into the intricate relationship between sex hormones and cancer, elucidating the underlying biological mechanisms and their clinical implications. We explore the multifaceted roles of estrogen, androgens, and progesterone, highlighting their respective influence on specific cancers such as breast, ovarian, endometrial, and prostate. Special attention is given to estrogen receptor-positive (ER+) and estrogen receptor-negative (ER-) tumors, androgen receptor signaling, and the dual role of progesterone in both promoting and inhibiting cancer progression. Clinical observations reveal varied treatment responses contingent upon hormonal levels, with certain therapies like tamoxifen, aromatase inhibitors, and anti-androgens demonstrating notable success. However, disparities in treatment outcomes between males and females in hormone-sensitive cancers necessitate further exploration. Therapeutically, the utilization of hormone replacement therapy (HRT) during cancer treatments presents both potential risks and benefits. The promise of personalized therapies, tailored to an individual's hormonal profile, offers a novel approach to optimizing therapeutic outcomes. Concurrently, the burgeoning exploration of new drugs and interventions targeting hormonal pathways heralds a future of more effective and precise treatments for hormone-sensitive cancers. This review underscores the pressing need for a deeper understanding of sex hormones in cancer therapy and the ensuing implications for future therapeutic innovations.

Clinicopathological features, psychological status, and prognosis of 33 patients with occult breast cancer.

Occult breast cancer (OBC) has traditionally been considered to be a carcinoma of unknown primary origin with a favorable prognosis and can be treated as stage II-III breast cancer. Due to the small number of cases and limited clinical ex-perience, treatments vary greatly around the world and no standardized treat-ment has yet been established. To investigate the clinicopathological features, psychological status and prog-nostic features of patients with OBC. The clinicopathological data of 33 OBC patients diagnosed and treated in the Affiliated Hospital of Xuzhou Medical University and Xuzhou Central Hospital from November 2015 to November 2022 were retrospectively analyzed. The psychological status of OBC patients was evaluated by the Self-rating Anxiety Scale and Self-rating Depression Scale. Patients' emotions, stress perception and psychological resilience were evaluated by the Positive and Negative Affect Schedule, the Chinese Perceived Stress Scale, and the Connor-Davidson Resilience Scale (CD-RISC), respectively. Patient survival was calculated using the Kaplan-Meier method, and survival curves were plotted for analysis with the log-rank test. Univariate and multivariate survival analyses were performed using the Cox regression model. The 33 OBC patients included 32 females and 1 male. Of the 33 patients, 30 (91%) had axillary tumors, 3 (9%) had a neck mass as the primary symptom; 18 (54.5%) had estrogen receptor-positive tumors, 17 (51.5%) had progesterone receptor-positive tumors, and 18 (54.5%) had Her-2-positive tumors; 24 (72.7%) received surgical treatment, including 18 patients who underwent modified radical mastectomy, 1 patient who underwent breast-conserving surgery plus axillary lymph node dissection (ALND), and 5 patients who underwent ALND alone; 12 patients received preoperative neoadjuvant therapy. All 30 patients developed anxiety and depression, with low positive affect scores and high negative affect scores, accompanied by a high stress level and poor psychological resilience. There were no differences in the psychological status of patients according to age, body mass index, or menopausal status. The overall survival and disease-free survival (DFS) of all the patients were 83.3% and 55.7%, respectively. Univariate analysis demonstrated that the initial tumor site (P = 0.021) and node stage (P = 0.020) were factors that may affect patient prognosis. The 5-year DFS rate of OBC patients who received radiotherapy was greater (P < 0.001), while the use of different surgical methods (P = 0.687) had no statistically significant effect on patient outcomes. Multivariate analysis revealed that radiotherapy (P = 0.031) was an independent prognostic factor. Receiving radiotherapy had a significant effect on the CD-RISC score (P = 0.02). OBC is a rare breast disease whose diagnosis and treatment are currently controversial. There was no significant difference in the efficacy of other less invasive surgical procedures compared to those of modified radical mastectomy. In addition, radiotherapy can significantly improve patient outcomes. We should pay attention to the psychological state of patients while they receive antitumor therapy.

The potential of epigenetic therapy to target the 3D epigenome in endocrine-resistant breast cancer

Three-dimensional (3D) epigenome remodeling is an important mechanism of gene deregulation in cancer. However, its potential as a target to counteract therapy resistance remains largely unaddressed. Here, we show that epigenetic therapy with decitabine (5-Aza-mC) suppresses tumor growth in xenograft models of pre-clinical metastatic estrogen receptor positive (ER+) breast tumor. Decitabine-induced genome-wide DNA hypomethylation results in large-scale 3D epigenome deregulation, including de-compaction of higher-order chromatin structure and loss of boundary insulation of topologically associated domains. Significant DNA hypomethylation associates with ectopic activation of ER-enhancers, gain in ER binding, creation of new 3D enhancer–promoter interactions and concordant up-regulation of ER-mediated transcription pathways. Importantly, long-term withdrawal of epigenetic therapy partially restores methylation at ER-enhancer elements, resulting in a loss of ectopic 3D enhancer–promoter interactions and associated gene repression. Our study illustrates the potential of epigenetic therapy to target ER+ endocrine-resistant breast cancer by DNA methylation-dependent rewiring of 3D chromatin interactions, which are associated with the suppression of tumor growth.

Uncovering a novel mechanism: Butyrate induces estrogen receptor alpha activation independent of estrogen stimulation in MCF-7 breast cancer cells.

Butyrate is a promising candidate for an antitumoral drug, as it promotes cancer cell apoptosis and reduces hormone receptor activity, while promoting differentiation and proliferation in normal cells. However, the effects of low-dose butyrate on breast cancer cell cultures are unclear. We explored the impact of sub-therapeutic doses of butyrate on estrogen receptor alpha (ERα) transcriptional activity in MCF-7 cells, using RT-qPCR, Western blot, wound-healing assays, and chromatin immunoprecipitation. Our results showed that sub-therapeutic doses of sodium butyrate (0.1 - 0.2 mM) increased the transcription of ESR1, TFF1, and CSTD genes, but did not affect ERα protein levels. Moreover, we observed an increase in cell migration in wound-healing assays. ChIP assays revealed that treatment with 0.1 mM of sodium butyrate resulted in estrogen-independent recruitment of ERα at the pS2 promoter and loss of NCoR. Appropriate therapeutic dosage of butyrate is essential to avoid potential adverse effects on patients' health, especially in the case of estrogen receptor-positive breast tumors. Sub-therapeutic doses of butyrate may induce undesirable cell processes, such as migration due to low-dose butyrate-mediated ERα activation. These findings shed light on the complex effects of butyrate in breast cancer and provide insights for research in the development of antitumoral drugs.

Unlocking the potential of aromatase inhibitors: recent advances in drug design, synthesis, docking activity, and in vitro bioactivity evaluations

Breast cancer, a global health concern claiming approximately 685,000 lives in 2020, necessitates continual advancements in therapeutic strategies. Estrogen and aromatase play pivotal roles in hormone-responsive breast cancer, with 80% of patients exhibiting estrogen receptor-positive tumors. Aromatase inhibitors (AIs), notably non-steroidal inhibitors like anastrozole and letrozole, have significantly improved outcomes, yet challenges persist, including side effects. This review focuses on recent developments in AIs, exploring xanthone derivatives, imidazole derivatives, and curcumin derivatives as potential inhibitors of aromatase. Molecular docking studies, employing Auto Dock and other tools, reveal the binding affinities and interactions of these compounds with the aromatase enzyme. Among xanthones, Erythrommone emerges as a potent inhibitor, holding promise for clinical trials. Imidazole derivatives, synthesized through the Debus-Radziszewski reaction, demonstrate anticancer potential, with compounds like 1a exhibiting superior efficacy against MCF7 cells. ADME-Tox analyses indicate promising drug-likeness but reveal potential mutagenic effects and environmental impacts. Curcumin derivatives, particularly 1,5-diaryl-1,4-pentadien-3-ones, present alternatives to address curcumin's bioavailability challenges. A study of 25 compounds (DKC) identifies DKC-10 as a potent inhibitor, outperforming established breast cancer drugs in terms of binding affinity and interactions with aromatase and ERα+ receptors. These findings underscore the importance of exploring diverse chemical structures in developing AIs, paving the way for more effective and well-tolerated therapeutics. The integration of computational techniques, such as molecular docking studies, accelerates drug discovery by predicting interactions at the molecular level. Overall, this comprehensive review provides valuable insights into the evolving landscape of aromatase inhibitors, offering a roadmap for future research and the development of advanced breast cancer therapeutics.

Radiometal Complexes as Pharmacokinetic Modifiers: A Potent 68Ga-Labeled Gastrin-Releasing Peptide Receptor Antagonist Based on the Macrocyclic Metal Chelator NODIA-Me.

The gastrin-releasing peptide receptor (GRPr) is overexpressed in various cancer types including prostate and breast carcinomas, making it an attractive target for molecular imaging and therapy. In this work, we designed a novel GRPr antagonistic probe comprising metal chelator NODIA-Me. This 1,4,7-triazacyclononane-based chelator forms positively charged metal complexes due to its neutral methylimidazole arms. Because a positive charge at the N-terminus of GRPr conjugates is responsible for high receptor affinity as exemplified by the current gold standard DOTA-RM2, we investigated if a positively charged radiometal complex can be used as a pharmacokinetic modifier to also produce high-affinity GRPr conjugates. In this respect, the bioconjugate NODIA-Me-Ahx-JMV594 was prepared by a combination of solid-phase peptide synthesis and solution-based reactions in a 94% yield. Radiolabeling provided the 68Ga-labeled conjugate in radiochemical yields of >95% and radiochemical purities of >98% with mean molar activities of Am ∼17 MBq nmol-1. The competitive GRPr affinity of the metal-free and 69/71Ga-labeled conjugate was determined to be IC50 = 0.41 ± 0.06 and 1.45 ± 0.06 nM, respectively. The metal-free GRPr antagonist DOTA-RM2 and its corresponding 69/71Ga complex had IC50 values of 1.42 ± 0.07 and 0.98 ± 0.19 nM, respectively. Small-animal PET imaging of mice bearing GRPr(+) PC-3 tumors revealed high radioactivity accumulation in the tumors and in the pancreas as an organ with high levels of GRPr expression. These findings were corroborated by the corresponding ex vivo biodistribution data, in which the tumors and the pancreas exhibited the highest radioactivity accumulation. By coinjection of an excess of NODIA-Me-Ahx-JMV594, uptake in the tumors and GRPr(+) organs was significantly reduced, confirming specific receptor-mediated uptake. The estrogen receptor-positive tumor of a female breast cancer patient was clearly visualized by PET imaging using 68Ga-labeled NODIA-Me-Ahx-JMV594. To summarize, the positive charge at the N-terminus of the conjugate induced by the Ga(NODIA-Me) complex resulted in high GRPr affinity comparable to that of the potent antagonist DOTA-RM2. The conjugate NODIA-Me-Ahx-JMV594 is a promising probe for imaging of GRPr tumors that warrants further evaluation in larger patient cohorts as well as in combination with other radiometals.

Statin Use, Cholesterol Level, and Mortality Among Females With Breast Cancer

Several studies have reported an association between the use of statins and breast cancer (BC) mortality. However, most of these studies did not take into account the underlying cholesterol level. To investigate the association between serum cholesterol, statin use, and BC mortality. This cohort study included females with invasive BC that was newly diagnosed between January 1, 1995, and December 31, 2013, in Finland. The cohort had available hormone receptor data and at least 1 cholesterol measurement. All data were obtained from Finnish national registries. Statistical analyses were performed from January to May 2022. Use of statins; statin dose; and serum cholesterol, low-density lipoprotein, high-density lipoprotein, and triglyceride levels measured separately before and after BC diagnosis. Breast cancer mortality and overall mortality between date of BC diagnosis and December 31, 2015. A total of 13 378 female patients with BC (median [IQR] age, 62 [54-69] years) participated in the study. The median (IQR) follow-up was 4.5 (2.4-9.8) years after BC diagnosis, during which 16.4% of patients died and 7.0% died of BC. Prediagnostic statin use was a risk factor for BC death even after adjustment for total cholesterol level (hazard ratio [HR], 1.22; 95% CI, 1.02-1.46; P = .03). Reduced risk for BC death was seen for postdiagnostic statin use (HR, 0.85; 95% CI, 0.73-1.00; P = .05). The risk reduction was robust in participants whose cholesterol level decreased after starting statins (HR, 0.49; 95% CI, 0.32-0.75; P = .001) but was nonsignificant if cholesterol level did not subsequently decrease (HR, 0.69; 95% CI, 0.34-1.40; P = .30). Reduced BC mortality among statin users was also observed in females with estrogen receptor-positive tumors (HR, 0.82; 95% CI, 0.68-0.99; P = .03). Overall mortality was lower among statin users vs nonusers when adjusted for serum cholesterol level (HR, 0.80; 95% CI, 0.72-0.88; P < .001). Results of this cohort study showed that postdiagnostic use of statins was associated with reduced BC mortality compared with nonuse, and the risk was associated with subsequent change in serum cholesterol level. This finding suggests that cholesterol-lowering interventions with statins may be beneficial for patients with BC.

Understanding the role of environmental and socioeconomic factors in the geographic variation of breast cancer risk in the US-wide Sister Study

ObjectiveTo describe the geographic pattern of breast cancer incidence in a nationwide prospective cohort and investigate whether environmental exposures and/or neighborhood socioeconomic status explain observed geographic disparities. MethodsUsing accelerated failure time models with a spatial random effect term, we mapped the health region-level association between residential location and breast cancer incidence for 44,707 participants in the Sister Study after controlling for established individual-level breast cancer risk factors. We performed a variable selection process to select environmental exposures [i.e., ambient nitrogen dioxide (NO2) and fine particulate matter (PM2.5), PM2.5 chemical composition, outdoor light at night (LAN), ambient noise, ultraviolet radiation, and greenspace] and neighborhood-level factors [i.e., population density and area deprivation index (ADI)] that predicted breast cancer incidence and quantified the spatial variation explained by the selected factors. We also considered whether the geographic pattern and predictors were similar when restricting to estrogen receptor-positive (ER+) tumors. ResultsWe observed a spatial patterning in the incidence of overall breast cancer (Moran's I = 16.7, p < 0.05) and ER+ breast cancer (Moran's I = 13.2, p < 0.05), with a lower risk observed in the South and Southeast and a greater risk in the Northwest and certain areas of the Midwest and Northeast. NO2, LAN, and ADI explained 21.4% of the spatial variation in overall breast cancer incidence whereas NO2, PM2.5 chemical composition, LAN, greenspace, and ADI together explained 63.3% of the spatial variation in ER+ breast cancer incidence. ConclusionsOur findings provide additional evidence for a role of environmental exposures in breast cancer incidence and suggest that geographic-based risk factors may vary according to breast cancer subtype. Our findings support the need for additional research to quantify the relative contributions of geographic-based risk factors for breast cancer.

THU570 Isoform Specific Gene Regulation By Progesterone Receptors Drives Expansion Of Breast Cancer Stem Cell Expansion In 3D Cell Culture Models

Abstract Disclosure: N.E. Gillis: None. T.H. Truong: None. C.A. Lange: None. Exposure to progesterone is a risk factor for breast cancer, and numerous PGR polymorphisms are linked to breast, endometrial, and ovarian cancer. Two progesterone receptor (PR) isoforms are expressed from PGR in breast tissues: full-length PR-B and truncated PR-A. In normal breast epithelia, PR isoforms are equally expressed and mostly function as A:B heterodimers. An imbalance of this ratio, and thus a predominance of homodimers, is frequent in ER+ breast cancer. Luminal A-type estrogen receptor positive (ER+) tumors often exhibit an elevated PR-A:PR-B ratio that predicts resistance to tamoxifen. It has recently been shown that PRs promote endocrine resistance and drive expansion of breast cancer stem-like cell (CSC) populations. However, the isoform-specific molecular mechanisms of PR action in ER+ breast cancers remain undefined. The goal of our current work is to map the progesterone-driven gene expression in CSC cells through transcriptomic profiling of T47D cells grown in spheroid-culture conditions that express exclusively PR-A or PR-B. This transcriptomic data has been integrated with phenotypic studies of T47D and MCF7 cells that express a single isoform of PR. Our data shows PR-A is a regulator of the cell cycle and of senescence, while PR-B mediates cell metabolism and intracellular signaling. We further integrated our cell line data with transcriptomes from patient tumors that have been classified based on their PR-A:PR-B ratio and their response to treatment with anti-progestins. Our results are a first step towards re-defining the role of PRs in gene regulation within the CSC population. Presentation Date: Thursday, June 15, 2023

Tumor Ki-67, ER and PR, and antibodies against estradiol and progesterone in breast cancer patients

Aim. To investigate the associations of cell proliferation marker Ki-67 in estrogen receptor (ER) and progesterone receptor (PR) positive (ER+/ PR+) and negative (ER-/PR-) tumors with the ratio of antibodies against estradiol and progesterone (IgA1-E2/IgA1-Pg) in the serum of breast cancer (BC) patients.Materials and Methods. Antibodies against steroid hormones were analyzed by ELISA in the serum of 432 healthy women and 1212 patients with BC (573 patients with I stage and 639 patients with II–IV stages). Expression of Ki-67, ER and PR in tumors was determined by immunohistochemical staining. Serum estradiol and progesterone were measured by enzyme-linked immunosorbent assay.Results. In total, low IgA1-E2/IgA1-Pg (≤ 1) and high IgA1-E2/IgA1-Pg (&gt; 1) ratio were revealed in 49.3% and 50.7% of healthy women; in 25.7% and 74.3% of stage I BC patients with tumor Ki-67 &lt; 14 (р &lt; 0.001; OR = 0.4 and OR = 2.8, respectively), and in 17.1% and 82.9% of stage I BC patients with tumor Ki-67 &gt; 30 (р &lt; 0.001; OR = 0.2 and OR = 4.7, respectively). The differences between patients with low and high tumor Ki-67 levels in relation to low and high IgA1-E2/IgA1-Pg ratio were statistically significant (p = 0.03). In stage I BC patients with ER+/PR+ and tumors with Ki-67 &lt; 14, low and high IgA1-E2/IgA1-Pg ratio were found in 25.0% and 75.0% cases (р &lt; 0.001; OR = 0.3 and OR = 2.9, respectively). In stage I BC patients with ER+/PR+ and tumors with Ki-67 &gt; 30, low and high IgA1-E2/IgA1-Pg ratio were found in 12.9% and 87.1% cases (р &lt; 0.001; OR = 0.2 and OR = 6.6, respectively). In patients with ER+/PR+ tumors, the differences between patients with low and high tumor Ki-67 levels in relation to low and high IgA1-E2/IgA1-Pg ratio were also statistically significant (p = 0.009). In patients with ER-/PR- tumors, the differences between patients with low and high Ki-67 levels in relation to low and high IgA1-E2/IgA1-Pg ratio were not revealed. The proportion of breast cancer patients with tumor Ki-67 &gt; 30 increased from I to II–IV BC stages regardless of IgA1-E2/IgA1-Pg ratio.Conclusion. IgA1-E2/IgA1-Pg ratio may serve as a predictor of tumor proliferative activity in stage I BC patients with ER+/PR+ tumors.