Treatment Of ER-positive Breast Cancer Research Articles

Arctigenin inhibits proliferation of ER-positive breast cancer cells through cell cycle arrest mediated by GSK3-dependent cyclin D1 degradation

Estrogen receptor (ER) positive accounts for a large proportion of breast cancer. Although there are many targeted therapeutic drugs, the emergence of drug resistance urgently requires the development of new drugs. Arctigenin (Arc), a lignan found in certain plants of the Asteraceae, has the effect on inhibiting breast cancer, but its molecular mechanism has not been clear. AimsTo this end, the current study focuses on understanding the mechanism of Arc on ER-positive breast cancer cells. Main methodsColony formation experiments and sulforhodamine B methods were used to determine the growth-inhibitory effect of Arc. The cell cycle and apoptosis were analyzed by flow cytometry. Alterations of signaling proteins were measured by Western blotting. Protein degradation was determined by comparing protein half-lives and inhibiting proteasome. Key findingsThe experimental results show that Arc did not induce apoptosis in ER-positive breast cancer cell, rather caused G1 cycle arrest by decreasing cyclin D1 levels without effect on altering CDK4/6 levels. Moreover, we have demonstrated that Arc decreases cyclin D1 levels through prompting Akt/GSK3β-mediated degradation. SignificanceThese findings warrant the potential of Arc as a candidate treatment for ER-positive breast cancer.

Taking a Second Look at Neoadjuvant Endocrine Therapy for the Treatment of Early Stage Estrogen Receptor Positive Breast Cancer During the COVID-19 Outbreak.

Taking a Second Look at Neoadjuvant Endocrine Therapy for the Treatment of Early Stage Estrogen Receptor Positive Breast Cancer During the COVID-19 Outbreak.

G1T48, an oral selective estrogen receptor degrader, and the CDK4/6 inhibitor lerociclib inhibit tumor growth in animal models of endocrine-resistant breast cancer

PurposeThe combination of targeting the CDK4/6 and estrogen receptor (ER) signaling pathways with palbociclib and fulvestrant is a proven therapeutic strategy for the treatment of ER-positive breast cancer. However, the poor physicochemical properties of fulvestrant require monthly intramuscular injections to patients, which limit the pharmacokinetic and pharmacodynamic activity of the compound. Therefore, an orally available compound that more rapidly reaches steady state may lead to a better clinical response in patients. Here, we report the identification of G1T48, a novel orally bioavailable, non-steroidal small molecule antagonist of ER.MethodsThe pharmacological effects and the antineoplastic mechanism of action of G1T48 on tumors was evaluated using human breast cancer cells (in vitro) and xenograft efficacy models (in vivo).ResultsG1T48 is a potent and efficacious inhibitor of estrogen-mediated transcription and proliferation in ER-positive breast cancer cells, similar to the pure antiestrogen fulvestrant. In addition, G1T48 can effectively suppress ER activity in multiple models of endocrine therapy resistance including those harboring ER mutations and growth factor activation. In vivo, G1T48 has robust antitumor activity in a model of estrogen-dependent breast cancer (MCF7) and significantly inhibited the growth of tamoxifen-resistant (TamR), long-term estrogen-deprived (LTED) and patient-derived xenograft tumors with an increased response being observed with the combination of G1T48 and the CDK4/6 inhibitor lerociclib.ConclusionsThese data show that G1T48 has the potential to be an efficacious oral antineoplastic agent in ER-positive breast cancer.

Mathematical modeling of ER-positive breast cancer treatment with AZD9496 and palbociclib

Breast cancer is a heterogeneous disease. Understanding the tumor subtypes can help identify the best option of treatment. Mathematical modeling of tumor growth is a useful tool to understand the observed clinical phenomena. This work is based on a previously published mathematical model of breast cancer in MCF-7 cell line. In this paper, cancer therapy using AZD9496 and palbociclib is incorporated into the mathematical model. Numerical results produced using the mathematical model are compared with those observed in clinical and experimental studies. Numerical simulation shows that monotherapy with AZD9496 can inhibit the growth of a tumor with a restricted tumor size, monotherapy with palbociclib is ineffective, and combination therapy with AZD9496 and palbociclib can produce synergistic effect and control a larger tumor.

Androgen Receptor Is a Non-canonical Inhibitor of Wild-Type and Mutant Estrogen Receptors in Hormone Receptor-Positive Breast Cancers.

SummarySustained treatment of estrogen receptor (ER)-positive breast cancer with ER-targeting drugs results in ER mutations and refractory unresponsive cancers. Androgen receptor (AR), which is expressed in 80%–95% of ER-positive breast cancers, could serve as an alternate therapeutic target. Although AR agonists were used in the past to treat breast cancer, their use is currently infrequent due to virilizing side effects. Discovery of tissue-selective AR modulators (SARMs) has renewed interest in using AR agonists to treat breast cancer. Using translational models, we show that AR agonist and SARM, but not antagonist, inhibit the proliferation and growth of ER-positive breast cancer cells, patient-derived tissues, and patient-derived xenografts (PDX). Ligand-activated AR inhibits wild-type and mutant ER activity by reprogramming the ER and FOXA1 cistrome and rendering tumor growth inhibition. These findings suggest that ligand-activated AR may function as a non-canonical inhibitor of ER and that AR agonists may offer a safe and effective treatment for ER-positive breast cancer.

Andrographolide is an Alternative Treatment to Overcome Resistance in ER-Positive Breast Cancer via Cholesterol Biosynthesis Pathway

Oestrogen receptor (ER)-positive breast cancer is one of the common forms of breast cancer affecting women worldwide. ER-positive breast cancer patients are subjected to anti-oestrogen therapy such as selective oestrogen receptor modulator (SERM) and aromatase inhibitors (AIs). Recently, the emergence of resistance to anti-oestrogen treatment is under intensive focus. The different mechanisms postulated to explain the occurrence of resistance in ER-positive breast cancer treatment include the loss of ER function and the crosstalk between signalling pathways in cancer cells. Recent literature highlighted that the cholesterol biosynthesis pathway acts as a novel mechanism underlying resistance to oestrogen deprivation. The present study aimed to highlight the role of cholesterol biosynthesis in anti-oestrogen treatment resistance, putatively suggesting an alternative plant-based treatment using andrographolide from Andrographis paniculata. The hypolipidaemic effect of andrographolide can be utilised to prevent the resistance in the treatment of ER-positive breast cancer contributed by cholesterol biosynthesis.

Development of a Liposomal Formulation of Acetyltanshinone IIA for Breast Cancer Therapy.

Acetyltanshinone IIA (ATA), synthesized in our group exhibiting good anti-breast cancer effects, is expected to replace the commonly used anti-ER+ breast cancer (breast cancer cells overexpressing the estrogen receptor) drug tamoxifen. To promote the clinical progress of ATA, polyethylene glycol (PEG)-modified liposomes were used to encapsulate ATA along with improving its bioavailability and in vivo anticancer efficiency. The resulting liposomal ATA exhibited a spherical shape with an average size of 188.5 nm. In vitro evaluations showed that liposomal ATA retained the anti-breast cancer efficacy of ATA while exerting much less cytotoxicity toward noncancerous cells. Significantly, pharmacokinetics analysis showed that the AUC0-24h of liposomal ATA was 59 times higher than that of free ATA, demonstrating increased bioavailability of ATA. Preclinical experiments demonstrated that liposomal ATA reduced the growth of ER-positive human breast tumor xenografts by 73% in nude mice, and the liposomal ATA exhibited a much lower level of toxicity than that of free ATA with respect to zebrafish larval mortality, body formation, and heart function during development. Moreover, 7-day and 21-day tissue toxicity levels were determined in mice by intravenous administration of a maximum dosage of liposomal ATA (120 mg/kg). The results showed no obvious tissue damage in major organs, including the heart, liver, spleen, kidney, and brain. In summary, we have developed a clinical formulation of liposomal ATA with the high bioavailability and potent efficacy for the treatment of ER-positive breast cancer.

Sequential versus concurrent use of chemotherapy and endocrine therapy in the adjuvant treatment of ER-positive breast cancer: A systematic review and Bayesian network meta-analysis.

e12040 Background: Chemotherapy followed by endocrine therapy is the standard adjuvant treatment strategy for estrogen receptor-positive breast cancer patients. However, no direct evidence so far demonstrated better efficacy of sequential use of chemotherapy and endocrine therapy over concurrent. Objective: To evaluate the efficacy between sequential and concurrent use of chemotherapy and endocrine therapy in the adjuvant treatment of ER positive breast cancer. Methods: Randomized clinical trials comparing chemotherapy and/or endocrine therapy in the adjuvant treatment of ER positive breast cancer were included. Hazard ratios (HRs) of disease-free survival (DFS) and overall survival (OS) were extracted and analyzed in Bayesian analysis. Patients were stratified by menopause status for subgroup analysis. Results: 37 trials were identified with 37225 patients enrolled in total, 37 trials with DFS results and 24 with OS. 3 comparisons were done between sequential and concurrent arms. In DFS analysis, no statistical significance was found in all 3 comparisons [CHE seq/con TAM (HR 1.01, 95%CI 0.8497 - 1.199); CHE seq/con OFS+TAM (HR 0.9119, 95%CI 0.5666 - 1.49); CHE seq/con OFS+AI (HR 1.032, 95%CI 0.6291 - 1.776)]. The same were seen in OS analysis [CHE seq/con TAM (HR 0.9512, 95%CI 0.8053 - 1.125); CHE seq/con OFS+TAM (HR 1.065, 95%CI 0.6344 - 1.789); CHE seq/con OFS+AI (HR 1.069, 95%CI 0.665 - 1.717)]. Rankings were done for preferable treatment recommendations. In DFS analyses, sequential arms ranked higher than concurrent arms [CHE seq/con OFS+AI (1 vs. 3); CHE seq/con OFS+TAM (6 vs. 7); CHE seq/con TAM (8 vs. 8)]. The same tendency was seen in OS analyses [CHE seq/con OFS+AI (1 vs. 2); CHE seq/con TAM (4 vs. 5)] except for CHE seq/con OFS+TAM (11 vs. 6-9). In subgroup ranking results, CHE seq/con OFS+AI and CHE seq/con OFS+TAM showed consistency among comparisons with concurrent arms ranked higher than sequential arms. However, CHE seq TAM ranked higher than CHE con TAM in all comparisons. Conclusions: The combination of chemotherapy and endocrine therapy in the adjuvant treatment of ER positive breast cancer demonstrated equal efficacy either used sequentially or concurrently. However, concurrent arms were recommended over sequential arms in premenopausal patients for better DFS and OS, except for the combination of chemotherapy and tamoxifen which was recommended to be used sequentially. Others: This study has been registered in PROSPERO (CRD42018104889).

Structure-activity relationship study of estrogen receptor down-regulators with a diphenylmethane skeleton.

Selective estrogen receptor (ER) down-regulators (SERDs) are pure ER antagonists that also induce ER degradation upon binding to the receptor. Although SERDs have been developed for the treatment of ER-positive breast cancers for nearly a decade, their precise mechanism(s) of action and structure-activity relationship are still unclear. Generally, Western blotting is used to examine the effects of SERDs on ER protein levels, but the methodology is low-throughput and not quantitative. Here, we describe a quantitative, high-throughput, luciferase-based assay for the evaluation of SERDs activity. For this purpose, we established stable recombinant HEK-293 cell lines expressing ERα fused with emerald luciferase. We also designed and synthesized new diphenylmethane derivatives as candidate SERDs, and evaluated their SERDs activity using the developed system in order to examine their structure-activity relationship, taking EC50 as a measure of potency, and Emax as a measure of efficacy.

Long noncoding RNA signature in predicting metastasis following tamoxifen treatment for ER-positive breast cancer.

We aimed to develop a long noncoding RNA (lncRNA) expression signature that can predict response to tamoxifen. LncRNA expression profiling was mined in two cohorts from Gene Expression Omnibus (GSE6532, GSE9195, n=412). A set of lncRNAs (LINC01191, RP4-639F20.1 and CTC-429P9.3) associated with distant metastasis-free survival was established. Estrogen receptor-positive breast cancer patients in the training series could be classified into high- and low-risk groups with significantly different distant metastasis-free survival values based on this signature (hazard ratio [HR]: 5.11; p=7.28×10-8). The prognostic ability of this signature was confirmed in validation sets 1 (HR: 2.58; p=1.54×10-2) and 2 (HR: 10.06; p=6.85×10-3). The lncRNA signature may have possible clinical implications in the selection of high-risk patients for tamoxifen therapy.

Abstract 1850: CDK inhibitor PHA-848125 preferentially inhibits proliferation of triple negative breast cancer and synergizes with cisplatin

Abstract Triple-negative breast cancer (TNBC), defined by the lack of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2), is highly aggressive with a poor prognosis as it lacks targeted therapies. Several inhibitors of the cyclin-dependent kinase 4 and 6 (CDK4/6) have been approved by the FDA for the treatment of ER-positive breast cancer. However, the effectiveness of these inhibitors against TNBC tumors is less than expected. Here, we tested the efficacy of PHA-848125, a potent ATP-competitive CDK2 inhibitor, on a panel of breast cancer cell lines showing a selective impaired proliferation in TNBC cells (IC50 of ER-positive MCF7 cells: 25.9 μM vs. IC50 of TNBC MDA-MB-231 cells: 430.8 nM). TNBC cells treated with PHA-848125 exhibited effects consistent with CDK2 inhibition, including G1 cell cycle arrest and reduced phosphorylation levels of the retinoblastoma protein (pRB) and CDK2. Interestingly, PHA-848125 was also effective against pRB-null TNBC cell lines, unlike other approved CDK inhibitors. Oral administration of PHA-848125 to TNBC xenograft mice and MMTV-PyMT transgenic mice impaired both tumor growth and metastasis at a tolerable dose (40 mg/kg), but not in ER-positive xenograft mice. A combination treatment of PHA-848125 and cisplatin inhibited tumor growth and induced apoptosis synergistically in vitro and in vivo. Thus, our preclinical experiments set the rationale for the clinical evaluation of PHA-848125, either alone or in combination with cisplatin, for the treatment of TNBC. Citation Format: Douglas G. Cheung, Marta Buzzetti, Carlo M. Croce, Gianpiero di Leva. CDK inhibitor PHA-848125 preferentially inhibits proliferation of triple negative breast cancer and synergizes with cisplatin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1850.

Efficacy and tolerability of AFPep, a cyclic peptide with anti-breast cancer properties

PurposeThe purpose of this study is to assess the efficacy and safety profile of AFPep, a 9-amino acid cyclic peptide prior to its entry into pre-clinical toxicology analyses en route to clinical trials. MethodsAFPep was assessed for anti-estrogenic activity in a mouse uterine growth assay and for breast cancer therapeutic efficacy in a human tumor xenograft model in mice. AFPep was assessed for tolerability in a variety of in vivo models, notably including assessment for effects on rat liver and human hepatocellular carcinoma cell lines and xenografts. ResultsAFPep arrests the growth of human MCF-7 breast cancer xenografts, inhibits the estrogen-induced growth of mouse uteri, and does not affect liver growth nor stimulate growth of human hepatocellular carcinoma cell lines when growing in vitro or as xenografts in vivo. AFPep is well tolerated in mice, rats, dogs, and primates. ConclusionsAFPep is effective for the treatment of ER-positive breast cancer and exhibits a therapeutic index that is substantially wider than that for drugs currently in clinical use. The data emphasize the importance of pursuing pre-clinical toxicology studies with the intent to enter clinical trials.

Bone and calcium metabolism associated with malignancy. Endocrine therapy of breast cancer.

Estrogen receptor(ER)-positive breast cancer accounts for approximately 80%of all breast cancers. The basic concept of endocrine therapy is to deplete estrogen(LH-RH agonist and aromatase inhibitor)and inhibit the action of estrogen against estrogen-dependent growth in ER-positive breast cancer. Recently, new molecular targeting agents, including an mTOR inhibitor and a CDK4/6 inhibitor, have been used in combination with endocrine therapy for the treatment of ER-positive metastatic breast cancer.

Abstract 474: Estrogen-responsive miRNAs as modulators of E2-induced apoptosis in AI-resistant breast cancer

Abstract Long-term estrogen deprivation with tamoxifen or aromatase inhibitors (AI) is the basic principle of endocrine treatment of ER-positive breast cancer. Acquired resistance is however a major obstacle in treatment success. Based on clinical observations and in vitro as well as animal in vivo experiments it has been suggested that tumor cell clones evolve over time and become vulnerable to E2-induced apoptosis, thereby potentially providing a promising second line treatment option. In vitro, this vulnerability is mimicked in the AI-resistant breast cancer models MCF-7:5C and MCF-7:2A, which during long-term E2-deprivation reconfigure their survival signaling including endoplasmic reticulum, oxidative and inflammatory stress related pathways. E2-stimulation of these cells initiates an unfolded protein response which in turn triggers apoptosis through the intrinsic and subsequently extrinsic pathway (Jordan VC 2015). Recently, we identified miRNA profiles matching the biology of AI resistance and vulnerability to E2-induced apoptosis (Hoppe R et al. 2016). Here we investigate the modulatory role of miRNAs in E2-induced apoptosis through the identification of their global expression changes in E2-stimulated 5C and 2A models compared to MCF-7:WS8 reference. Each cell line was treated with 10-9 M E2 or vehicle over a 72h time course (6, 12, 24, 72 h). miRNA profiles were generated using Affymetrix GeneChip miRNA2.0 arrays. At each time point relative miRNA expression changes (E2/control) were evaluated resulting in a total of 72 (5C), 104 (2A) and 94 (WS8) differentially expressed miRNAs (FC > 1.5 or < 1/1.5, Pt-test < 0.05). Differential expression analyses between consecutive time points revealed 16 (5C), 43 (2A) and 27 (WS8) miRNAs (maximum absolute FC difference > 1.5, PF-test < 0.05). Differential area under the curve (dAUC) analyses at 6-72, 6-24, and 24-72 h mined the overall, early, and late-responding miRNAs of the different phenotypes (Ppermutation test < 0.05). Representative early down-regulated miRNA candidates potentially modulating E2-induced apoptosis in 5C cells are miR-543 and miR-432 of the DLK1-DIO3 locus on Chr. 14q32.31. Late responding up-regulated miRNA candidates are miR-150* and miR-149* the low expression of which in tumor tissues registered in The Cancer Genome Atlas (miRNA-Seq v. 3.1.17.0) was associated with worse outcome in all PAM50 breast cancer subtypes and Lum B, respectively (HR = 2.1, 95% CI: 1.4-3.2; P = 0.00017; HR = 6.8, 95% CI: 1.5-3.0; P = 0.0036). We are currently in the process to subject respective miRNA sets to functional enrichment analyses (KEGG, GO) and to correlate E2-responsive miRNA candidates with respective transcriptome signatures. In summary, we will present miRNAs matching to the biological processes inherent to E2-induced apoptosis. These may serve as potential targets for the amplification of the apoptotic trigger upon E2-treatment. Citation Format: Reiner Hoppe, Ping Fan, Stefan Winter, Florian Büttner, V. Craig Jordan, Hiltrud B. Brauch. Estrogen-responsive miRNAs as modulators of E2-induced apoptosis in AI-resistant breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 474. doi:10.1158/1538-7445.AM2017-474

A Review of Fulvestrant in Breast Cancer

Fulvestrant is a selective estrogen receptor degrader that binds, blocks and degrades the estrogen receptor (ER), leading to complete inhibition of estrogen signaling through the ER. This review article further explains the mechanism of action of the drug and goes on to review the trials carried out to optimize its dosing. Multiple trials have been undertaken to compare fulvestrant with other endocrine treatments, and results have shown it to have similar efficacy to anastrozole, tamoxifen and exemestane at 250 mg every 28 days. However, when given at 500 mg every 28 days, with an extra loading dose on day 14, it has demonstrated an improved progression-free survival (PFS) compared to anastrozole. We look at how fulvestrant has been used in combination with CDK4/6 inhibitors such as palbociclib (PALOMA-3) and ribociclib (MONALEESA-3) and drugs targeting the PI3K/AKT/mTOR pathway such as pictilisib (FERGI) and buparlisib (BELLE-2 and BELLE-3). We then go on to describe a selection of the ongoing clinical trials looking at combination therapy involving fulvestrant. Finally, we review the effect of fulvestrant in patients who have developed resistance to aromatase inhibitors via ESR1 mutation, where it has been shown to offer a PFS benefit that is further improved by the addition of the CDK4/6 inhibitor palbociclib. Whilst fulvestrant is clearly an effective drug as monotherapy, we believe that its role in the treatment of ER-positive breast cancer may be best reserved for combination therapy, and whilst there are multiple trials currently in progress, it would appear that the combination with CDK4/6 inhibitors would offer the greatest promise in terms of balancing benefit with toxicity.

Down-regulation of Forkhead box protein A1 (FOXA1) leads to cancer stem cell-like properties in tamoxifen-resistant breast cancer cells through induction of interleukin-6

The selective estrogen receptor (ER) modulator tamoxifen inhibits ER signaling in breast cancer cells, and it is used for the treatment of ER-positive breast cancer. However, this type of cancer often acquires resistance to tamoxifen, and a better understanding of the molecular mechanisms underlying tamoxifen resistance is required. In this study, we established tamoxifen-resistant (TAM-R) breast cancer cells by long-term tamoxifen treatment of ER-positive breast cancer MCF7 cells. In TAM-R cells, expression of not only ERα, a major form of ER in breast cancer, but also its transcriptional partner forkhead box protein A1 (FOXA1) was found to be reduced. In contrast, activation of the transcription factor nuclear factor-κB (NF-κB) and expression of its target IL6 were increased in these cells. Stable expression of FOXA1, but not ERα, reduced the expression of IL6 in the FOXA1- and ERα-negative breast cancer MDA-MB-231 cells and TAM-R cells, without affecting the activation of the NF-κB signaling pathways. Conversely, FOXA1 knockdown induced IL6 expression in MCF7 cells. Chromatin immunoprecipitation assays revealed that FOXA1 bound to the promoter region of IL6 and repressed recruitment of the NF-κB complex to this region. TAM-R cells were found to have high mammosphere-forming activity, characteristics of cancer stem cells, and this activity was suppressed by NF-κB and IL6 signaling inhibitors. Taken together, these results suggest that FOXA1 suppresses expression of IL6 through inhibition of NF-κB recruitment to the IL6 promoter in an ERα-independent manner and that reduction in FOXA1 expression induces IL6 expression and contributes to cancer stem cell-like properties in TAM-R cells.

Hormone-Targeted Therapy and Resistance

It has been 40 years since the US Food and Drug Administration approved the estrogen receptor (ER) antagonist tamoxifen for the treatment of ER-positive breast cancer, ushering in the era of targeted therapy coupled with a companion diagnostic. The prostate cancer field quickly followed suit with the approval of the androgen receptor (AR) antagonist bicalutamide. In the years since, there has been sustained scientific interest in understanding these hormone-dependent signaling pathways and in drug discovery efforts to identify novel hormone-directed therapeutic agents. Recently, there have been breakthrough discoveries relating to mechanisms that enable reactivation of ER and AR signaling in the presence of antihormonal agents and that enable loss of hormone dependency, providing multiple routes of acquired resistance to hormone therapy. This review discusses parallels between breast and prostate cancer, including their pathobiologies, existing therapeutic modalities, acquired resistance to such therapeutics, and novel therapies being developed to target distinct states of resistance.

Abstract P3-04-22: RAD1901 demonstrates anti-tumor activity in multiple models of ER-positive breast cancer treatment resistance

Abstract Estrogen receptor positive (ER+) breast cancer makes up approximately 70% of all newly diagnosed breast cancers and ER remains the single most useful predictive and prognostic marker for these patients. In fact, dependence on ER signaling is retained in recurrent disease and even in disease treated with multiple lines of anti-endocrine agents. Indeed, inhibiting estrogen synthesis (e.g. with aromatase inhibitors) and modulating ER pathway activity (e.g. with tamoxifen) continue to be mainstays in the standard of care for ER+ breast cancer patients both in adjuvant and metastatic setting. While patients initially respond well to these agents, a majority of patients with metastatic disease will relapse. Therefore, it is critical to improve our understanding and model the specific mechanisms that can lead to treatment resistance. Mutations in the ligand binding domain of ESR1 have been described as a mechanism that contributes to clinical resistance to aromatase inhibitors. In addition, some ESR1 mutations can lead to a conformational change of the receptor, leading to decreased binding of tamoxifen, thereby reducing its activity. Recently, selective estrogen receptor degraders, or SERDs, have gained widespread attention as a novel treatment strategy for ER+ disease. Indeed, fulvestrant has been shown to degrade ER and cause tumor growth inhibition in many ER+ breast cancer models. However, in the clinic, fulvestrant appears to be limited by PK exposure properties and this, combined with its intramuscular route of administration underscore the need for an oral SERD with improved PK properties. Recently, we have described RAD1901, a novel, orally available, non-steroidal small molecule SERD that is able to degrade ER and effectively cause tumor growth inhibition in multiple patient-derived xenograft models (PDX) models, including two that harbor a Y537S mutation in ESR1. To further validate that RAD1901 has the potential to be used in the advanced disease setting, we tested its anti-tumor activity in preclinical models representing the most common ESR1 mutations (D538G, E380Q, S463P, Y537N/C). In addition, we have tested RAD1901 in multiple cell line models of anti-endocrine resistance. These include long-term estrogen deprived cells that model progression of disease on aromatase inhibitors, as well as tamoxifen-resistant models. Here, we present anti-tumor activity of RAD1901 both as a single agent and as part of rational clinical combinations (eg. CDK4/6, PI3K/Akt/mTOR inhibitors) in multiple models of anti-endocrine resistance. These results combined with previous studies demonstrating RAD1901 efficacy in treatment naïve models provide strong preclinical evidence for clinical testing of RAD1901 in both adjuvant and metastatic setting. RAD1901 is currently under clinical investigation for the treatment of ER+ breast cancers in post-menopausal women (NCT02650817, NCT02338349). Citation Format: Bihani T, Arlt H, Brechbuhl H, Patel H, Tao N, Purandare DM, Hattersley G, Kabos P, Garner F. RAD1901 demonstrates anti-tumor activity in multiple models of ER-positive breast cancer treatment resistance [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P3-04-22.

Inhibition of Nicotinamide Phosphoribosyltransferase Induces Apoptosis in Estrogen Receptor-Positive MCF-7 Breast Cancer Cells.

PurposeTumor cells have increased turnover of nicotinamide adenine dinucleotide (NAD+), the main coenzyme in processes including adenosine diphosphate-ribosylation, deacetylation, and calcium mobilization. NAD+ is predominantly synthesized in human cells via the salvage pathway, with the first component being nicotinamide. Nicotinamide phosphoribosyltransferase (NAMPT) is the key enzyme in this pathway, and its chemical inhibition by FK866 has elicited antitumor effects in several preclinical models of solid and hematologic cancers. However, its efficacy in estrogen receptor (ER)-positive and human epidermal growth factor receptor 2-positive breast cancer cells has not been previously investigated. In this study, we aimed to deplete the NAD+ content of MCF-7 cells, a model cell line for ER-positive breast cancer, by inhibiting NAMPT in order to evaluate downstream effects on p53 and its acetylation, p21 and Bcl-2-associated X protein (BAX) expression, and finally, apoptosis in MCF-7 breast cancer cells.MethodsMCF-7 cells were cultured and treated with FK866. NAD+ levels in cells were determined colorimetrically. Levels of p53 and its acetylated form were determined by Western blotting. Expression of p21 and BAX was determined by real-time polymerase chain reaction. Finally, levels of apoptosis were assessed by flow cytometry using markers for annexin V and propidium iodide.ResultsFK866 treatment was able to increase p53 levels and acetylation, upregulate BAX and p21 expression, and induce apoptosis in MCF-7 cells. Addition of exogenous NAD+ to cells reversed these effects, suggesting that FK866 exerted its effects by depleting NAD+ levels.ConclusionResults showed that FK866 could effectively inhibit NAD+ biosynthesis and induce programmed cell death in MCF-7 cells, suggesting that NAMPT inhibitors may be useful for the treatment of ER-positive breast cancers.

Anti-miR-203 suppresses ER-positive breast cancer growth and stemness by targeting SOCS3.

Breast cancer is a major public health problem worldwide in women and existing treatments are not adequately effective for this deadly disease. microRNAs (miRNAs) regulate the expression of many target genes and play pivotal roles in the development, as well as in the suppression of many cancers including breast cancer. We previously observed that miR-203 was highly upregulated in breast cancer tissues and in ER-positive breast cancer cell lines. In our present study, we observed that anti-miR-203 suppresses breast cancer cell proliferation in vitro. Orthotopic implantation of miR-203 depleted MCF-7 cells into nude mice displays smaller tumor growth as compared to control MCF-7 cells. Furthermore, miR-203 expression is significantly higher in ER-positive breast cancer patients as compared to ER-negative patients. We identified suppressor of cytokine signaling 3 (SOCS3) as a direct target of miR-203. Here we observed that miR-203 expression is inversely correlated with SOCS3 expression in ER-positive breast cancer samples. Additionally, we found that anti-miR-203 suppressed the expression of pStat3, pERK and c-Myc in MCF-7 and ZR-75-1 cells. We also demonstrated that anti-miR-203 decreased mammospheres formation and expression of stem cell markers in MCF-7 and ZR-75-1 cells. Taken together, our data suggest that anti-miR-203 has potential as a novel therapeutic strategy in ER-positive breast cancer treatment.