Estrogen Receptor Signaling Pathways Research Articles

Interaction of Bisphenol A and Its Analogs with Estrogen and Androgen Receptor from Atlantic Cod (Gadus morhua).

The widespread use of bisphenol A (BPA) in polycarbonate plastics and epoxy resins has made it a prevalent environmental pollutant in aquatic ecosystems. BPA poses a significant threat to marine and freshwater wildlife due to its documented endocrine-disrupting effects on various species. Manufacturers are increasingly turning to other bisphenol compounds as supposedly safer alternatives. In this study, we employed in vitro reporter gene assays and ex vivo precision-cut liver slices from Atlantic cod (Gadus morhua) to investigate whether BPA and 11 BPA analogs exhibit estrogenic, antiestrogenic, androgenic, or antiandrogenic effects by influencing estrogen or androgen receptor signaling pathways. Most bisphenols, including BPA, displayed estrogenic properties by activating the Atlantic cod estrogen receptor alpha (gmEra). BPB, BPE, and BPF exhibited efficacy similar to or higher than that of BPA, with BPB and BPAF being more potent agonists. Additionally, some bisphenols, like BPG, induced estrogenic effects in ex vivo liver slices despite not activating gmEra in vitro, suggesting structural modifications by hepatic biotransformation enzymes. While only BPC2 and BPAF activated the Atlantic cod androgen receptor alpha (gmAra), several bisphenols exhibited antiandrogenic effects by inhibiting gmAra activity. This study underscores the endocrine-disrupting impact of bisphenols on aquatic organisms, emphasizing that substitutes for BPA may pose equal or greater risks to both the environment and human health.

A phase 2, open-label, single-arm, prospective, multicenter study of nab-sirolimus plus letrozole in advanced or recurrent endometrioid endometrial cancer.

TPS5640 Background: This clinical trial is designed to evaluate the antitumor activity of nab-sirolimus in combination with letrozole in patients with advanced or recurrent endometrioid endometrial carcinoma (EEC). Despite recent pivotal data demonstrating improved outcomes with immunotherapy plus chemotherapy, regardless of mismatch repair status, alternative nonchemotherapy-based treatment options for patients with advanced or recurrent EEC remain necessary. Dysregulation of mTOR signaling is implicated in the pathology of EEC, in which >80% harbor PTEN or PI3K/Akt/mTOR pathway alterations. Moreover, crosstalk between mTOR and estrogen receptor signaling pathways is associated with endocrine therapy resistance. GOG-3007 and other phase 2 studies have demonstrated that the combination of conventional mTOR inhibitors (mTORis) and endocrine therapy provides clinical benefit in patients with EEC. nab-Sirolimus is an mTOR inhibitor that uses albumin-bound sirolimus nanoparticles given by IV and is approved by the US FDA for malignant perivascular epithelioid cell tumor. Preclinical data of nab-sirolimus demonstrated improved tumor accumulation, mTOR inhibition, and tumor growth suppression compared with conventional mTORis. We hypothesize that the simultaneous inhibition of mTOR and ER with nab-sirolimus and letrozole, respectively, may provide a signal of synergistic antitumor activity in patients with recurrent EEC. Methods: In this phase 2, open-label, single-arm, multicenter study (NCT05997017), nab-sirolimus (100 mg/m2, intravenous, on days 1 and 8 of each 21-day cycle) and letrozole (2.5 mg, oral, daily) are administered to patients (~29 planned, Simon’s 2-stage design) with clinically confirmed, advanced, or recurrent EEC. Eligibility criteria include age ≥18 years, ≤1 prior chemotherapy-based regimen in the advanced, metastatic, or recurrent setting, prior adjuvant chemotherapy is permitted, Eastern Cooperative Oncology Group performance status score 0 or 1, naive to mTORi, and measurable disease (per Response Evaluation Criteria in Solid Tumors [RECIST] v1.1). The primary endpoint is best overall response rate by RECIST v1.1. Key secondary endpoints include duration of response, disease control rate (defined as complete response plus partial response plus stable disease of ≥12 weeks after start of treatment), progression-free survival, overall survival, and safety. Exploratory endpoints include correlation of baseline molecular biomarkers with clinical outcomes. The trial is open for enrollment. Clinical trial information: NCT05997017 .

Developmental Delay and Male-Biased Sex Ratio in esr2b Knockout Zebrafish.

The estrogen receptor signaling pathway plays an important role in vertebrate embryonic development and sexual differentiation. There are four major estrogen receptors in zebrafish: esr1, esr2a, esr2b and gper. However, the specific role of different estrogen receptors in zebrafish is not clear. To investigate the role of esr2b in zebrafish development and reproduction, this study utilized TALENs technology to generate an esr2b knockout homozygous zebrafish line. The number of eggs laid by esr2b knockout female zebrafish did not differ significantly from that of wild zebrafish. The embryonic development process of wild-type and esr2b knockout zebrafish was observed, revealing a significant developmental delay in the esr2b knockout zebrafish. Additionally, mortality rates were significantly higher in esr2b knockout zebrafish than in their wild-type counterparts at 24 hpf. The reciprocal cross experiment between esr2b knockout zebrafish and wild-type zebrafish revealed that the absence of esr2b resulted in a decline in the quality of zebrafish oocytes, while having no impact on sperm cells. The knockout of esr2b also led to an abnormal sex ratio in the adult zebrafish population, with a female-to-male ratio of approximately 1:7. The quantitative PCR (qPCR) and in situ hybridization results demonstrated a significant downregulation of cyp19ab1b expression in esr2b knockout embryos compared to wild-type embryos throughout development (at 2 dpf, 3 dpf and 4 dpf). Additionally, the estrogen-mediated induction expression of cyp19ab1b was attenuated, while the estradiol-induced upregulated expression of vtg1 was disrupted. These results suggest that esr2b is involved in regulating zebrafish oocyte development and sex differentiation.

Royal jelly and its hormonal effects in breast cancer: a literature review.

Breast cancer is the most common cancer in women which can be cured in most individuals with early-stage non-metastatic disease. Imbalance in estrogen signaling pathways and propagating levels of estrogens has important roles in breast cancer development. Targeting the estrogen receptor signaling pathway is linked to breast cancer treatment. Royal jelly is one of the bee products containing 10-hydroxy-2-decenoic acid, a structure similar to mammalian estrogen, allowing it to attach to estrogen receptors. It is considered as a general tonic and immunomodulator which may be helpful in reducing the side effects of cancer treatments. Currently, there are controversial data regarding the pros and cons of royal jelly in cancer. Here we provide an overview of the effects of royal jelly on sex hormones and its possible role in breast cancer. Electronic databases including PubMed, Scopus, and Web of Science were searched with the search terms royal jelly, cancer, and sexual hormones. All preclinical and clinical studies regarding the hormonal effects of royal jelly were included. According to the collected preclinical data, consumption of royal jelly at daily doses below 200mg/kg can be useful to decrease the risk of breast cancer since it reduces the serum level of estrogen; whereas increases progesterone, which subsequently decreases the expression of ERs on the ER-positive cells. Future clinical studies are essential to confirm the safe dose of royal jelly as an adjuvant therapy in breast cancer.

Abstract PO2-01-13: Clinico-Pathological and Molecular Characterization of Early Hormone Receptor-Positive Breast Cancer in Young Women

Abstract Introduction Premenopausal women diagnosed with hormone receptor-positive (HR+)/HER2-negative (HER2-) early breast cancer (EBC) face a significantly higher risk of cancer recurrence and mortality compared to older women, particularly those diagnosed under the age of 40. The underlying causes behind this disparity remain largely unknown. Material and methods We retrospectively select tumour samples from patients (pts) diagnosed with HR+/HER2- EBC at Hospital Clinic (Barcelona, Spain) from May 2014 to March 2020 who had a PAM50/Prosigna assay (Veracyte) previously performed. This assay examines the expression of 50 genes involved in important breast cancer signaling pathways, as well as the intrinsic subtypes (IS) and the Risk of Recurrence (ROR) score. Our primary objective was to compare the distinct clinicopathological and molecular characteristics of HR+/HER2- EBC between two age groups: patients under 40 years old (Young EBC or YEBC) and those 40 years old and above (Older EBC or OEBC). Disease-free survival (DFS) was defined as the time between diagnosis and the first recurrence. Comparisons between groups were carried out using chi-square tests (categorical variables). Additionally, we performed a two-class unpaired Significance Analysis of Microarrays (SAM) with a false discovery rate (FDR) less than 5% to identify genes with significantly different expression between YEBC and OEBC. The statistical significance level was set at less than 0.05. Results A total of 441 tumor samples from 420 patients were included in the analysis, with 5% (22/441) corresponding to YEBC. With a median follow-up of 64.5 months, there were 20 breast cancer (BC) recurrences (4.8%), 12 second cancers (2.9%) and 5 BC-related deaths (1.2%), all occurring in the OEBC group. Table 1 provides an overview of the main clinico-pathological and molecular characteristics. YEBC tended to have larger tumors, with 45.5% (10/22) measuring over 2 cm compared to 36.1% (151/419) in OEBC (p=0.376). YEBC also had a higher proportion of grade 3 tumors (18.18% versus 3.1% in OEBC, p=0.418) and lower expression of estrogen receptor (ER) as determined by immunohistochemistry (mean of 72.28% vs 88.36%, p=0.004). Regarding the PAM50 IS, there was a difference observed between YEBC and OEBC (p < 0.001). Specifically, YEBC patients exhibited a tendency towards a higher proportion of Luminal B tumors (54.6% versus 38.9% in OEBC, p=0.144), as well as non-Luminal subtypes such as Basal-like (18.2% versus 0.95% in OEBC, p< 0.001) and HER2E (4.6% versus 0.7% in OEBC, p=0.186). YEBC tended to have higher ROR (p=0.167), and to receive chemotherapy in a higher proportion, especially in the intermediate-risk group (p=0.004). In YEBC tumours, SAM analysis (p < 0.05) showed an overexpression of genes mainly related with proliferation (i.e., RRM2, MELK, MIK67), cell cycle progression (i.e., KNTC2, CDC20, MYBL2) and apoptosis inhibition (i.e., BIRC5). In OEBC there were an overexpression of genes related with the oestrogen receptor-signalling pathway (i.e., ESR1, NAT1, FOXA1, PGR). Luminal B tumours in YEBC showed an upregulation of genes related to proliferation, migration, and poor prognosis (i.e., KRT17, KRT5) compared to OEBC. However, since there were no recurrences in the YEBC group, differences in terms of DFS between the groups could not be calculated. Principio del formulario Final del formulario Conclusions YEBC exhibited a more advanced stage and a more aggressive intrinsic phenotype compared to OEBC. Additionally, these tumors showed a tendency to upregulate genes involved in proliferation, cell cycle progression, and survival, while downregulating genes related to hormone signaling pathways. These molecular characteristics may help explain the worse prognosis of YEBC compared to OEBC. However, further research is necessary to delve into the differential genomic characteristics that contribute to the unfavorable prognosis observed in patients diagnosed with HR+ EBC before the age of 40. Key clinical and molecular features in 441 biopsies Citation Format: Francisco Javier Muñoz-Carrillo, José Antonio Sola, Carme Crous, Esther Sanfeliu, Isabel Garcia-Fructuoso, Elia Seguí, Teresa Gorría, Raquel Gómez-Bravo, Barbara Adamo, Tomás Pascual, Olga Martínez-Sáez, Francesco Schettini, Pablo Rivera, Rosalía Cayuela, Montserrat Muñoz, Núria Chic, Maria Vidal. Clinico-Pathological and Molecular Characterization of Early Hormone Receptor-Positive Breast Cancer in Young Women [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 PO2-01-13.

Abstract RF02-01: A Multicenter, Phase I/II Trial of Anastrozole, Palbociclib, Trastuzumab, and Pertuzumab in Hormone Receptor (HR)-Positive, HER2-Positive Metastatic Breast Cancer (ASPIRE)

Abstract Background: Among patients (pts) with HR-positive and HER2-positive breast cancer, crosstalk between HER2 and estrogen receptor (ER) signaling pathways may contribute to endocrine resistance but anti-HER2 agents in combination with endocrine therapy can restore endocrine sensitivity. Mechanistically, HER2/HER3 signaling promotes survival by way of PI3K-AKT activation whereas ER-CDK4/6-Rb signaling promotes cell cycle progression. The combination of anti-HER2 therapy with an aromatase inhibitor (AI) and a CDK 4/6 inhibitor would allow for blockade of both pathways and provide a novel, all biologic, and chemotherapy-free approach to the treatment of HR-positive, HER2-positive metastatic breast cancer (MBC). Methods: We conducted a phase I/II multi-institution trial in pts with previously untreated, HR-positive and HER2-positive MBC. In the Phase I portion, pts received escalating doses of palbociclib (100mg, 125mg) in conjunction with trastuzumab, pertuzumab and an AI anastrozole, using a 3+3 dose escalation trial design. In the phase II portion, pts received palbociclib at the maximum tolerated dose (MTD), anastrozole, trastuzumab, and pertuzumab. The primary endpoints of the Phase I and II portions were MTD and clinical benefit rate (CBR) defined as the sum of complete response, partial response, and stable disease for >/= 6 months, respectively. Secondary endpoints included progression free survival (PFS), objective response rate (ORR) and safety. The Phase II portion of this study was powered with 30 pts to show efficacy of palbociclib administered at the MTD in combination with anastrozole, trastuzumab and pertuzumab if the CBR achieved at 6 months exceeded 58%. The Clopper-Pearson method was used to calculate confidence intervals for ORR and CBR. The PFS distribution was estimated using the Kaplan-Meier method. Results: In the Phase I portion, a total of 9 pts were enrolled. No DLTs were observed at the 100mg dose (N=3) or the 125mg dose (N=6) level, and thus, 125mg was established as the MTD. An additional 24 pts were enrolled to the Phase II portion at the MTD, with a total of 30 pts in the modified intention-to-treat population included in the efficacy analysis. The median age of the population was 57.6 years (range 50.5-63.8) and 27% of pts were premenopausal and received ovarian function suppression as part of treatment. As shown in Table 1, the primary endpoint, CBR, was 97% (95% CI: 0.83-1.0, p<0.0001). ORR was 70% (95% CI: 0.51-0.85). Median time to objective response was 2.8 months with earliest response at 3 months. Median duration of response was not reached with range of 5.1-42.2 months. Median PFS was also not reached with range of 8-44.8 months. Safety data were consistent with known toxicity profiles of agents. Most common adverse events included diarrhea (80%), neutropenia (77%), leukopenia (70%), anemia (67%), and fatigue (60%). Grade 3-4 events occurred in 63% (19/30) of pts and included neutropenia (68%), leukopenia (32%), decrease in absolute neutrophil count (21%), and anemia (21%). Conclusions: The combination of anastrozole, palbociclib, trastuzumab, and pertuzumab was well tolerated and effective with a clinical benefit rate of 97% in pts with previously untreated HR-positive, HER2-positive MBC. The combination provides a chemotherapy-free alternative for pts with triple positive breast cancers. Further follow up will determine impact on PFS. Table 1. Results in Patients on Anastrozole, Palbociclib, Trastuzumab, and Pertuzumab Citation Format: Rima Patel, Krystal Cascetta, Paula Klein, Erin Moshier, Maryann Kwa, Julie Fasano, Anupama Goel, Melissa Accordino, Charles Shapiro, Rita Vaccaro, Gargi Atul Joshi, Joseph Sparano, Amy Tiersten. A Multicenter, Phase I/II Trial of Anastrozole, Palbociclib, Trastuzumab, and Pertuzumab in Hormone Receptor (HR)-Positive, HER2-Positive Metastatic Breast Cancer (ASPIRE) [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 RF02-01.

Abstract PO2-23-11: SMAD4 depletion contributes to endocrine therapy resistance by ERBB signaling in HR+HER2- breast cancer

Abstract Objectives: Endocrine therapy resistance is a significant clinical challenge for patients with estrogen receptor (ER)-positive breast cancer. Dysregulation of the ER and ERBB signaling pathways plays a key role in endocrine therapy resistance. However, it is unclear how these pathways are integrated during resistance development. SMAD4 is involved in multiple stages of tumorigenesis, but its role in endocrine resistance development remains elusive. Here, we aimed to investigate the role of SMAD4 in the development of acquired endocrine therapy resistance in ER-positive breast cancer. Methods: CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) screening was conducted to identify genes involved in regulating the sensitivity of T47D cells to 4-hydroxytamoxifen (OHT). Bioinformatics analysis was performed to explore the clinical significance of SMAD4. The differential expression of SMAD4 in cells before and after endocrine treatment was assessed by RT-qPCR and immunoblotting. Loss-/gain-of-function assays were conducted to validate the phenotype. RNA-seq and phenotype rescue experiments were used to explore the underlying mechanisms. Drug combination experiments were performed to assess the therapeutic effect on SMAD4-depleted cells. Results: CRISPR screening identified SMAD4 as a key determinant of endocrine therapy resistance. Bioinformatic analysis showed that SMAD4 expression was downregulated in breast cancer tissues and that low expression was associated with poor patient prognosis. Differential expression analysis after endocrine therapy treatment showed that endocrine therapy downregulated the expression of SMAD4. In vitro and in vivo models further demonstrated that SMAD4 downregulation leads to endocrine therapy resistance. Transcriptome sequencing analysis identified that the ER, ERBB and PI3K/Akt/mTOR signaling pathways were aberrantly activated upon SMAD4 depletion. Further analysis revealed that the PI3K/Akt/mTOR pathway may contribute to the regulation of ERBB on ER signaling to some extent. The aberrant activation of autophagy in phenotype rescue experiments was found to reduce the rescue effect of the combination of 4-hydroxytamoxifen (OHT) and lapatinib (LAPA). Finally, drug combination experiments verified that the combined use of OHT, LAPA and hydroxychloroquine (CQ) produced a synergistic effect in SMAD4-depleted cells. Conclusions: Taken together, our findings demonstrate that SMAD4 plays a crucial role in endocrine therapy resistance and suggest a reasonable treatment strategy for ER-positive breast cancer patients who are resistant to endocrine therapy. Citation Format: Kang Li, Dan Shu, Han Li. SMAD4 depletion contributes to endocrine therapy resistance by ERBB signaling in HR+HER2- breast cancer [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 PO2-23-11.

Proteomic profiling points to the significance of adipocyte-derived extracellular vesicles in diet-induced obesity

Extracellular vesicles (EVs) are lipid-bilayer membrane-enclosed vesicles well known to mediate cell-cell communication to maintain normal physiological functions. The diverse cargo of EVs encompassing proteins, lipids and miRNAs represent potential biomarkers and therapeutic targets. However, the role of adipocyte-derived EVs (AdEVs) in obesity is still unknown. To address this, we performed proteomic profiling of AdEVs isolated from the C57BL/6 male mice that were fed either a normal chow diet (NCD) or a high-fat high-sucrose diet (HFHSD) for 12 weeks starting at 4 weeks of age. AdEVs were obtained, through ultracentrifugation, from the conditioned media of cultured adipocytes derived from gonadal adipose tissue. Nanoparticle tracking analysis (NTA) of AdEVs showed that adipocytes from mice fed HFHSD tended to release more AdEVs despite the comparable size of AdEVs relative to those of mice fed NCD. Subsequently, proteins were isolated, digested and labeled with TMTpro16-plex for proteomic analysis. Raw data were processed in Scaffold, and a general bioinformatic and functional analysis conducted in Ingenuity Pathway Analysis Qiagen (IPA) ( https://digitalinsights.qiagen.com/explore/ga-ipa?utm_source=QDI_GA_SA_IPA_1122_PF&cmpid=QDI_GA_IPA ) and Srplot identified over 1100 proteins in AdEVs (protein threshold: 99%, minim peptide:2, peptide threshold: 95%, 0.00% FDR). Of these, 58 proteins were identified as upregulated while 41 proteins were downregulated in HFHSD-fed mice (Fold Change > 2, adjusted p < 0.05). With IPA analysis, the top 3 influenced pathways including mitochondrial dysfunction (27%), Sirtuin signaling pathway (22%), and Granzyme A signaling (38%) are predicted to be activated while the oxidative phosphorylation (47%), estrogen receptor signaling (15%) and neutrophil extracellular trap signaling pathway (14%) are predicted to be inhibited. The Gene Oncology (GO) functional enrichment analysis in Srplot revealed the involvement of identified differentially expressed proteins in various biological processes such as lipid metabolism, catabolic process processes, and cellular respiration. Experiments are underway to validate specific proteins involved in these biological processes. Furthermore, analysis of cellular components highlighted three notable terms including myelin sheath, collagen-containing extracellular matrix and organelle inner membrane. In conclusion, the present study elucidates the significance of AdEVs in the context of diet-induced obesity and underscores the potential role of mitochondrial dysfunction in the initiation and progression of obesity and its associated diseases. NIH Veterans Affairs. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Triclocarban and triclosan promote breast cancer progression in vitro and in vivo via activating G protein-coupled estrogen receptor signaling pathways

Triclocarban (TCC) and triclosan (TCS) have been detected ubiquitously in human body and evoked increasing concerns. This study aimed to reveal the induction risks of TCC and TCS on triple negative breast cancer through non-genomic GPER-mediated signaling pathways. Molecular simulation indicated that TCC exhibited higher GPER binding affinity than TCS theoretically. Calcium mobilization assay displayed that TCC/TCS activated GPER signaling pathway with the lowest observed effective concentrations (LOEC) of 10 nM/100 nM. TCC and TCS also upregulated MMP-2/9, EGFR, MAPK3 but downregulated MAPK8 via GPER-mediated signaling pathway. Proliferation assay showed that TCC/TCS induced 4 T1 breast cancer cells proliferation with the LOEC of 100 nM/1000 nM. Wound-healing and transwell assays showed that TCC/TCS promoted 4 T1 cells migration in a concentration-dependent manner with the LOEC of 10 nM. The effects of TCC on breast cancer cells proliferation and migration were stronger than TCS and both were regulated by GPER. TCC/TCS induced migratory effects were more significantly than proliferative effect. Mechanism study showed that TCC/TCS downregulated the expression of epithelial marker (E-cadherin) but upregulated mesenchymal markers (snail and N-cadherin), which was reversed by GPER inhibitor G15. These biomarkers results indicated that TCC/TCS-induced 4 T1 cells migration was a classic epithelial to mesenchymal transition mechanism regulated by GPER signaling pathway. Orthotopic tumor model verified that TCC promoted breast cancer in-situ tumor growth and distal tissue metastasis via GPER-mediated signaling pathway at human-exposure level of 10 mg/kg/d. TCC-induced tissue metastasis of breast cancer was more significantly than in-situ tumor growth. Overall, we demonstrated for the first time that TCC/TCS could activate the GPER signaling pathways to induce breast cancer progression.

The prognostic value of MEK pathway-associated estrogen receptor signaling activity for female cancers.

Other than for breast cancer, endocrine therapy has not been highly effective for gynecologic cancers. Endocrine therapy resistance in estrogen receptor positive gynecologic cancers is still poorly understood. In this retrospective study, we examined the estrogen receptor (ER) signaling pathway activities of breast, ovarian, endometrial, and cervical cancers to identify those that may predict endocrine therapy responsiveness. Clinical and genomic data of women with breast and gynecological cancers were downloaded from cBioPortal for Cancer Genomics. Estrogen receptor alpha (ESR1) expression level and sample-level pathway enrichment scores (EERES) were calculated to classify patients into four groups (low/high ESR1 and low/high EERES). Correlation between ESR1/EERES score and survival was further validated with RNAseq data from low-grade serous ovarian cancer. Pathway analyses were performed among different ESR1/EERES groups to identify genes that correlate with endocrine resistance, which are validated using Cancer Cell Line Encyclopedia gene expression and Genomics of Drug Sensitivity in Cancer data. We identified a novel combined prognostic value of ESR1 expression and the corresponding estrogen response signaling (EERES score) for breast cancer. The combined prognostic value (ESR1/EERES) may be applicable to other gynecologic cancers. More importantly, we discovered that ER signaling can cross-regulate MEK pathway activation. We identified downstream genes in the MEK pathway (EPHA2, INAVA, MALL, MPZL2, PCDH1, and TNFRSF21) that are potential endocrine therapy response biomarkers. This study demonstrated that targeting both the ER and the ER signaling activity related MEK pathway may aid the development of endocrine therapy strategies for personalized medicine.

Abstract 7196: Evaluation of nab-sirolimus in combination with fulvestrant or PI3K pathway inhibitors to overcome resistance in breast cancer cell lines

Abstract Background: Endocrine therapy (ET) is the standard of care for hormone receptor positive (HR+) breast cancer (BC), but ET resistance develops in most patients (pts). One resistance mechanism involves cross talk between the PI3K-AKT-mTOR and estrogen receptor (ER) signaling pathways, resulting in increased ER signaling and tumor proliferation. This is mitigated by mTOR inhibition as demonstrated by improved outcomes with exemestane plus the mTOR inhibitor (mTORi) everolimus in pts with HR+ advanced BC. While PI3K inhibitors (PI3Ki [e.g. alpelisib]) are efficacious in pts with HR+, PIK3CA-mutated BC, resistance can occur due to acquired genomic alterations in the PI3K-AKT-mTOR pathway. Combination therapy that targets a critical downstream node, mTOR, could delay or overcome acquired resistance due to activation of upstream components of the PI3K-AKT-mTOR pathway. nab-Sirolimus is an intravenously administered, albumin-bound nanoparticle form of the mTORi sirolimus, with significantly greater tumor accumulation, mTOR target suppression, and improved antitumor effects than oral mTORis in nonclinical tumor models. We evaluated the antiproliferative and cytotoxic effects of nab-sirolimus in combination with ET or PI3Kis in BC cell lines. Methods: HR+, PI3K-mutated BC cell lines (MCF7 or MDA-MB-361) were incubated for 5 days with increasing concentrations of nab-sirolimus and the selective ER degrader fulvestrant. MDA-MB-361 cells or HR-negative (HR−), PI3K-mutated BC cells, MDA-MD-453, were incubated for 5 days with increasing concentrations of nab-sirolimus and PI3K-AKT-mTOR pathway inhibitors gedatolisib or alpelisib. Antiproliferative/cytotoxic effects of single-agent and combination treatment were assessed via an automated trypan blue exclusion assay. Results: Addition of 20 or 80 nM nab-sirolimus to fulvestrant (12.5-500 nM) markedly decreased cell viability and nearly doubled cell death in MCF7 and MDA-MB-361 cells. Antiproliferative effects of low-dose gedatolisib (2.5-10 nM) were enhanced by 61-71% when nab-sirolimus (20 or 80 nM) was added to MDA-MB-453 cells. While low doses (2.5, 5, and 10 nM) of gedatolisib alone led to limited cytotoxicity (4.2-9.6%), up to 48.9% of cell death occurred when 20 or 80 nM nab-sirolimus was added. Results were similar with alpelisib (1 uM) plus nab-sirolimus (20 or 80 nM). Western blot showed increased p4EBP1 in response to fulvestrant or PI3Ki treatment, which was reversed with the addition of nab-sirolimus. Conclusions: nab-Sirolimus enhanced the cytotoxic effects of fulvestrant in HR+ BC cells and of PI3Kis in HR+ and HR− BC cells. Addition of nab-sirolimus to ET or other PI3K-AKT-mTOR pathway inhibitors may overcome mechanisms of resistance; further exploration is warranted to elucidate clinical relevance. Citation Format: Khine Nyein Myint, Sean Wallace, Shihe Hou, Maria Zalath, Brian McMorran, Andrew Kwon, Igor Vivanco. Evaluation of nab-sirolimus in combination with fulvestrant or PI3K pathway inhibitors to overcome resistance in breast cancer cell lines [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 7196.

Abstract A018: A phase 2, open-label, single-arm, prospective, multicenter study of nab-sirolimus plus letrozole in advanced or recurrent endometrioid endometrial cancer

Abstract Introduction/Objective: This trial in progress is designed to evaluate the anti-tumor activity of nab-sirolimus in combination with letrozole in patients with advanced or recurrent endometrioid endometrial carcinoma (EEC). Despite recent pivotal data demonstrating improved outcomes with immunotherapy plus chemotherapy, regardless of mismatch repair status, alternative treatment options, particularly for advanced or recurrent EEC remain necessary. Dysregulation of mammalian target of rapamycin (mTOR) signaling is implicated in the pathology of EEC, in which >80% harbor PTEN or PI3K/AKT/mTOR pathway alterations. Moreover, crosstalk between mTOR and estrogen receptor signaling pathways is associated with endocrine resistance. GOG-3007 and other phase 2 studies have demonstrated that the combination of conventional mTOR inhibitors (mTORis) and endocrine therapy provides clinical benefit in patients with EC. nab-Sirolimus, a nanoparticle injectable form of mTORi, is approved in the USA for malignant perivascular epithelioid cell tumor. Preclinical data of nab-sirolimus demonstrated improved tumor accumulation, mTOR inhibition, and tumor growth suppression compared with conventional mTORis. We hypothesize that nab-sirolimus in combination with letrozole may produce synergistic antitumor activity in patients with recurrent EEC. Methods: In this phase 2, open-label, single-arm, multicenter study (NCT05997017), nab-sirolimus (100 mg/m2, intravenous, on days 1 and 8 of each 21-day cycle) and letrozole (2.5 mg, oral, daily) are administered to patients (~29 planned) with clinically confirmed, advanced, or recurrent EEC. Eligibility criteria include age ≥18 years, ≤1 prior chemo-based regimen, Eastern Cooperative Oncology Group performance status score 0 or 1, naïve to mTORi, and measurable disease (per Response Evaluation Criteria in Solid Tumors [RECIST] v1.1). The primary endpoint is best overall response rate by RECIST v1.1. Key secondary endpoints include duration of response, progression-free survival, overall survival, and safety. The relationship between biomarkers and response outcomes is an exploratory endpoint. Citation Format: Lauren E. Dockery, Anna Priebe, Linda Duska, Angela K. Green, Cara Mathews, Fernanda Musa, David O'Malley, Allison Puechl, Li Ding, Anita N. Schmid, Willis H. Navarro, Brian Slomovitz, Kathleen Moore. A phase 2, open-label, single-arm, prospective, multicenter study of nab-sirolimus plus letrozole in advanced or recurrent endometrioid endometrial cancer [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 A018.

Cuproptosis-related genes predict prognosis and trastuzumab therapeutic response in HER2-positive breast cancer

Breast cancer is the most common diagnosed cancer, the HER2-positive subtype account for 15% of all breast cancer. HER2-targeted therapy is the mainstay treatment for HER2-positive breast cancer. Cuproptosis is a novel form of programmed cell death, and is caused by mitochondrial lipoylation and destabilization of iron-sulfur proteins triggered by copper, which was considered as a key player in various biological processes. However, the roles of cuproptosis-related genes in HER2-positive breast cancer remain largely unknown. In the present study, we constructed a prognostic prediction model of HER2-positive breast cancer patients using TCGA database. Dysregulated genes for cells resistant to HER2-targeted therapy were analyzed in the GEO dataset. KEGG pathway, GO enrichment and GSEA was performed respectively. The immune landscape of DLAT was analyzed by CIBERSORT algorithm and TIDE algorithm. HER2-positive breast cancer patients with high CRGs risk score showed shorter OS. DLAT was downregulated and correlated with better survival of HER2-positive breast cancer patients (HR = 3.30, p = 0.022). High expressed DLAT was associated with resistant to HER2-targeted therapy. Knocking down DLAT with siRNA increased sensitivity of breast cancer to trastuzumab. KEGG pathway and GO enrichment of DEGs indicated that DLAT participates in various pathways correlated with organelle fission, chromosome segregation, nuclear division, hormone-mediated signaling pathway, regulation of intracellular estrogen receptor signaling pathway, condensed chromosome and PPAR signaling pathway. There was a negative correlation between TIDE and DLAT expression (r = − 0.292, p < 0.001), which means high DLAT expression is an indicator of sensitivity to immunotherapy. In conclusion, our study constructed a four CRGs signature prognostic prediction model and identified DLAT as an independent prognostic factor and associated with resistant to HER2-targeted therapy for HER2-positive breast cancer patients.

Therapeutic implications of the interplay between interferons and ER in breast cancer

The involvement of interferons (IFNs) in various diseases, including breast cancer, has sparked controversy due to their diverse roles in immunity and significant impact on pathological mechanisms. In the context of breast cancer, the heightened expression of endogenous IFNs has been linked to anti-tumor activity and a favorable prognosis for patients. Within the tumor tissue and microenvironment, IFNs initiate a cascade of molecular events involving numerous factors, which can lead to either cooperative or repressive interactions. The specific functions of IFNs in breast cancer vary depending on the two major disease phenotypes: hormone dependent (or responsive) and hormone independent (or unresponsive) breast cancer. Hormone dependence is determined by the presence of estrogen receptors (ERs). The interplay between the IFN and ER signaling pathways, and the involvement of intermediate factors such as NFκB, are areas that have been somewhat under-researched, but that hold potential importance for the understanding and treatment of breast cancer. This review aims to provide a comprehensive overview of the actions of IFNs in breast cancer, particularly in relation to the different breast cancer phenotypes and the significance of comprehending the underlying mechanisms. Furthermore, the use of IFN-based therapies in cancer treatment remains a topic of debate and has not yet gained widespread acceptance. However, emerging discoveries may redirect focus towards the potential of IFN-based therapies.

A novel scaffold long-acting selective estrogen receptor antagonist and degrader with superior preclinical profile against ER+ breast cancer

Breast cancer is one of the most common malignant tumors in women worldwide, with the majority of cases showing expression of estrogen receptors (ERs). Although drugs targeting ER have significantly improved survival rates in ER-positive patients, drug resistance remains an unmet clinical need. Fulvestrant, which overcomes selective estrogen receptor modulator (SERM) and AI (aromatase inhibitor) resistance, is currently the only long-acting selective estrogen receptor degrader (SERD) approved for both first and second-line settings. However, it fails to achieve satisfactory efficacy due to its poor solubility. Therefore, we designed and synthesized a series of novel scaffold (THC) derivatives, identifying their activities as ER antagonists and degraders. G-5b, the optimal compound, exhibited binding, antagonistic, degradation or anti-proliferative activities comparable to fulvestrant in ER+ wild type and mutants breast cancer cells. Notably, G-5b showed considerably improved stability and solubility. Research into the underlying mechanism indicated that G-5b engaged the proteasome pathway to degrade ER, subsequently inhibiting the ER signaling pathway and leading to the induction of apoptosis and cell cycle arrest events. Furthermore, G-5b displayed superior in vivo pharmacokinetics and pharmacodynamics properties, coupled with a favorable safety profile in the MCF-7 tamoxifen-resistant (MCF-7/TR) tumor xenograft model. Collectively, G-5b has emerged as a highly promising lead compound, offering potent antagonistic and degradation activities, positioning it as a novel long-acting SERD worthy of further refinement and optimization.

RelB-activated GPX4 inhibits ferroptosis and confers tamoxifen resistance in breast cancer

Tamoxifen (TAM) resistance remains a major obstacle in the treatment of advanced breast cancer (BCa). In addition to the competitive inhibition of the estrogen receptor (ER) signaling pathway, damping of mitochondrial function by increasing reactive oxygen species (ROS) is critical for enhancing TAM pharmacodynamics. Here, we showed that RelB contributes to TAM resistance by inhibiting TAM-provoked ferroptosis. TAM-induced ROS level promoted ferroptosis in TAM-sensitive cells, but the effect was alleviated in TAM-resistant cells with high constitutive levels of RelB. Mechanistically, RelB inhibited ferroptosis by transcriptional upregulating glutathione peroxidase 4 (GPX4). Consequently, elevating RelB and GPX4 in sensitive cells increased TAM resistance, and conversely, depriving RelB and GPX4 in resistant cells decreased TAM resistance. Furthermore, suppression of RelB transcriptional activation resensitized TAM-resistant cells by enhancing ferroptosis in vitro and in vivo. The inactivation of GPX4 in TAM-resistant cells consistently resensitized TAM by increasing ferroptosis-mediated cell death. Together, this study uncovered that inhibition of ferroptosis contributes to TAM resistance of BCa via RelB-upregulated GPX4.

Comprehensive assessment of the estrogenic activity of resin composites

Resin-based dental composites have been developed to restore decayed teeth or modify tooth color due to their excellent physical and chemical properties. Such composites may have intrinsic toxicity due to components released into the mouth during the early stage of polymerization, and afterward as a result of erosion or material decomposition. In addition, resin-based dental composites have potential environmental pollutant by elution of monomers and degradation. Since certain monomers of resin matrices are synthesized from bisphenol A (BPA), which acts as an estrogenic endocrine disruptor, these resin matrices may have estrogenic activity. Therefore, the estrogenic endocrine-disrupting activity of various dental composites should be evaluated. In this study, we evaluated the estrogenic endocrine-disrupting activity of 10 resin composites by using a BRET-based estrogen receptor (ER)α and ERβ dimerization assays and ER transactivation assay. BPA, BisDMA, BisGMA, BisEMA, TEGDMA, HMBP, and DMPA mediated ERα dimerization, and BPA, BisDMA, and DMPA also mediated ERβ dimerization. Except for UDMA and CQ, all the compounds were identified as estrogen agonists or antagonists. In-depth information for the safe use of dental composites was acquired, and it was confirmed how the component of dental composites acts in the ER signaling pathway. Further studies on the low-dose and long-term release of these compounds are needed to ensure the safe use of these resin-based dental composites.

A Basic Review on Estrogen Receptor Signaling Pathways in Breast Cancer.

Breast cancer is the most common cancer and the deadliest among women worldwide. Estrogen signaling is closely associated with hormone-dependent breast cancer (estrogen and progesterone receptor positive), which accounts for two-thirds of tumors. Hormone therapy using antiestrogens is the gold standard, but resistance to these treatments invariably occurs through various biological mechanisms, such as changes in estrogen receptor activity, mutations in the ESR1 gene, aberrant activation of the PI3K pathway or cell cycle dysregulations. All these factors have led to the development of new therapies, such as selective estrogen receptor degraders (SERDs), or combination therapies with cyclin-dependent kinases (CDK) 4/6 or PI3K inhibitors. Therefore, understanding the estrogen pathway is essential for the treatment and new drug development of hormone-dependent cancers. This mini-review summarizes current literature on the signalization, mechanisms of action and clinical implications of estrogen receptors in breast cancer.

Abstract 1238: Loss of HSPD1 induces cellular plasticity in triple-negative breast cancer cells

Abstract Abstract Background Unlike Breast cancer (BRCA) luminal subtypes, Triple negative breast cancer (TNBC) subtype lacks expression of estrogen receptor-α (ERα/ESR1), progesterone receptor (PGR) and/or epidermal growth factor (EGF) family receptor HER2/ERBB2 where patients do not benefit from endocrine and HER2 based therapies in addition to its poor differentiation that results in greater aggressiveness and poorer prognosis. The 60 KD Heat shock protein (HSP60) is a Mitochondrial protein that plays a role in stabilizing and refolding proteins and is regulated by HSPD1 gene. In normal cells, HSPD1 gene expression is controlled, however in TNBC, HSPD1 overexpression is common and is associated with poor cancer-specific survival. This study aims to understand downstream signaling pathways dependent on the function of HSPD1 in TNBC. Method To understand downstream signaling pathways dependent on the function of HSPD1, we analyzed the transcriptomic profiles of metastatic MDA-MB-231-WT and HSPD1-KD cells. Results Bioinformatics data analysis shows that HSPD1 is overexpressed in TNBC, particularly in basal subtype. High expression of HSPD1 related to low survival in breast cancer patients. HSPD1 KD in MDA-MB-231 cells identified several differentially regulated genes (DEG) as compared to WT MDA-MB-231 cells by RNA-seq analysis. Pathway analysis of DEG showed downregulation of mitochondrial respiration, thermogenesis, citric acid cycle, PD-1 signaling while upregulation of estrogen dependent gene expression, estrogen mediated signaling, nuclear receptor transcription pathway. These suggests that HSP60 overexpression in TNBC promotes basal type phenotype. Downregulation of HSP60 expression upregulated hormone receptors, particularly estrogen receptor (ESR1) and androgen receptor (AR) expression, thus inducing basal to luminal switch in MDA-MB-231 cells. These findings suggest that loss of HSPD1 significantly Induces plasticity in TNBC. Conclusion Our results suggest that loss of HSPDI in the context TNBC cells may promotes oncogenic transformation to an intermediary luminal progenitor phenotype that gains differentiation plasticity based on the upregulation of Estrogen-receptors signaling pathways. Ongoing analyses will be performed to identify the possible links between loss of HSPD1 and key transcription factors that define mammary epithelial cell differentiation. Citation Format: Muhammad G. Khodary, Alehegne W. Yirsaw, Temesgen Samuel, Clayton Yates, Deepa Bedi. Loss of HSPD1 induces cellular plasticity in triple-negative breast cancer cells [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 1238.

Role of RUNX2 in breast cancer development and drug resistance (Review).

Breast cancer is the most common malignancy and ranks second among the causes of tumor-associated death in females. The recurrence and drug resistance of breast cancer are intractable due to the presence of breast cancer stem cells (BCSCs), which are adequate to initiate tumor formation and refractory to conventional remedies. Runt-related transcription factor 2 (RUNX2), a pivotal transcription factor in mammary gland and bone development, has also been related to metastatic cancer and BCSCs. State-of-the-art research has indicated the retention of RUNX2 expression in a more invasive subtype of breast cancer, and in particular, triple-negative breast cancer development and drug resistance are associated with estrogen receptor signaling pathways. The present review mainly focused on the latest updates on RUNX2 in BCSCs and their roles in breast cancer progression and drug resistance, providing insight that may aid the development of RUNX2-based diagnostics and treatments for breast cancer in clinical practice.