Tamoxifen In Patients Research Articles

Abstract P1-12-17: Mature data (> 25 years) on 2 years adjuvant tamoxifen treatment in premenopausal women with breast cancer: Time to re-emphasize the progesterone receptor as predictive factor?

Abstract Background In 2011 The Early Breast Cancer Trialists' Collaborative Group (EBCTCG) reported that five years of adjuvant tamoxifen (TAM) significantly reduces the 15-year risks of breast cancer recurrence and death. Moreover, the estrogen receptor (ER) was considered the clinically most relevant predictive factor for efficacy of TAM treatment. In premenopausal patients participating in a randomized trial of 2 years of adjuvant TAM, we have previously reported on increased recurrence free survival in patients with ER- and PR-positive disease and a trend for increased overall survival in patients with PR-positive tumors. The median follow-up time was 13.9 years for patients without events. Since the effect on mortality may increase beyond year 10, we investigated more mature of data. Aims To investigate the long-term effect of 2 years adjuvant TAM in premenopausal patients choosing cumulative mortality (CM) and cumulative breast cancer mortality (CBCM) as end-points. Methods Premenopausal patients (n=564) with stage II breast cancer were included in a Swedish randomized multicenter trial between 1986-1991 and allocated to 2 years of TAM (n=276) or no treatment (n=288). The hormone receptor status of the tumor was known for 541 (96%) of the patients included, determined by immunohistochemistry or cytosol-based methods. Less than 2% were treated with adjuvant chemotherapy and they were equally distributed between the two groups. Median follow-up time was 26.3 years (22.7-29.7). Mortality data was obtained from the Swedish Cause of Death Register. Results Death from any cause was recorded among 312 (55%) of the patients, and 265 (47%) of the deaths were due to breast cancer. Two years of TAM decreased CM and CBCM in all patients irrespective of hormonal receptor status (n=564) (CM: HR 0.81; 95% CI: 0.65-1.02, p=0.070; CBCM: HR 0.79; 95% CI: 0.61-1.01, p=0.058), as well as in patients with ER-positive disease (n=346) (CM: HR 0.77; 95% CI: 0.57-1.03, p=0.077; CBCM: HR 0.72; 95% CI: 0.52-1.01, p=0.051), however not strictly significant. Importantly, in patients with PR-positive tumors, TAM significantly reduced CM and CBCM (CM: HR 0.73; 95% CI: 0.55-0.98, p=0.037; CBCM: HR 0.68; 95% CI: 0.49-0.94, p=0.020). Moreover, in patients with ER- and PR positive tumors TAM decreased CM (HR 0.74; 95% CI: 0.55-1.01, p=0.057) and CBCM (HR 0.69; 95% CI: 0-49-0.97, p=0.033), whereas there was no effect of TAM in patients with ER-positive and PR-negative tumors (p=0.97 and p=0.99, respectively). Conclusions The present study demonstrates that 2 years of adjuvant TAM as monotherapy significantly reduced CM and CBCM in premenopausal women with PR-positive tumors as well as CBCM in ER- and PR-positive tumors, after > 25 years of follow-up. Five years of adjuvant TAM have been proven to reduce mortality rates at 15 years, but we herein show that also 2 years of TAM yields a survival benefit at 25 years. In the latest meta-analysis from EBCTCG, PR did not add predictive information in patients with ER-positive tumors, but according to our results the significance of PR, as a predictive factor for TAM efficacy, should be re-emphasized in premenopausal women. Citation Format: Maria Ekholm, Pär-Ola Bendahl, Mårten Fernö, Bo Nordenskjöld, Olle Stål, Lisa Rydén. Mature data (> 25 years) on 2 years adjuvant tamoxifen treatment in premenopausal women with breast cancer: Time to re-emphasize the progesterone receptor as predictive factor? [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P1-12-17.

Abstract P1-12-07: Characteristics of recurrence after completing adjuvant tamoxifen therapy for 5 years

Abstract Background Treatment with tamoxifen (TMX) reduces the recurrence rate and increase overall survival in patients with hormone receptor positive breast cancer. Up to now, 5-year TMX therapy is generally accepted, but it is demonstrated that the rate of late recurrence after 5 years is considerably higher in hormone receptor positive type than in other subtype. Several clinical trials such as ATLAS and aTTom showed the benefit of continuing tamoxifen up to 10 years instead of stopping at 5 years without increasing mortality due to the effect of extended tamoxifen medication. Method We collected data of 1633 hormone receptor positive breast cancer patients who received surgery at Seoul National University Hospital from 1997 to 2007, and had completed 5-year TMX therapy with no recurrence within 5 years after diagnosis. Mean age of the patients was 43.3, and the patients have estrogen receptor or progesterone receptor. We included from the stage I to stage IV patients underwent curative surgery and received adequate adjuvant therapy such as chemotherapy or radiation therapy after surgery. We excluded the cases treated aromatase inhibitor (AI) or switched to AI. Result Among these patients, recurrences after 5 years of TMX therapy were found in 93 patients (late recurrence group). Local recurrences and distant metastases were found in 43 and 50 patients, respectively. Electronic medical records were retrospectively reviewed for clinicopathological factors. When comparing between patients with no recurrence and patients with late recurrence, p53 and HER-2 expression were significantly related to late recurrence (p=0.01, p<0.001 respectively). Also when subgroup analysis was done for distant metastasis of late recurrence group, distant metastasis was significantly associated with HER-2 expression and high nuclear grade (p=0.005, p=0.006 respectively). There are no relation between late recurrence and age, stage and ki-67. Conclusion our data shows that p53 and HER-2 expression is associated to late recurrence and especially HER-2 expression is related to distant metastasis after completing TMX for 5 years. On the basis of the result of large clinical trials, extending TMX therapy significantly reduces recurrence rate and increase survival. Our result support continuing TMX in patients with HER-2 expression and high nuclear grade is considerable after 5 years of TMX medication. Citation Format: Eunshin Lee, Han-Byoel Lee, Young Joon Kang, Yun-Gyoung Kim, Tae-kyung Yoo, Jongjin Kim, Min Kyoon Kim, Hyeong-Gon Moon, Dong-Young Noh, Wonshik Han. Characteristics of recurrence after completing adjuvant tamoxifen therapy for 5 years [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P1-12-07.

Abstract P2-05-13: Transcriptomic profiling of patient sequential tumours provides cutting edge view of global metastatic expression changes following endocrine therapy

Abstract Disease recurrence is a common problem in breast cancer and yet the mechanisms enabling tumour cells to evade therapy and colonise distant organs remain unclear. Here, for the first time, RNAsequencing has been performed on matched primary, nodal and liver metastatic tumours from three tamoxifen-treated patients following metastatic disease progression. Despite all primary tumours being of a luminal subtype and all cancers metastasising to the liver, the extent of patient heterogeneity at the gene level was striking. Less than 3% of the genes differentially expressed between sequential tumours were common to all patients. Larger divergence was observed between primary and liver tumours than between primary and nodal tumours, reflecting both the latency time to disease progression and the genetic impact of endocrine therapy. Furthermore, a xenograft model demonstrated the ability of tamoxifen to drive disease progression and establish distant metastatic disease in the endocrine resistant setting. Common functional pathways altered during metastatic, endocrine-resistant progression included ECM receptor interactions and focal adhesions. This novel global analysis highlights the influence of primary tumour biology in determining the transcriptomic profile of metastatic tumours, as well as the need for adaptations in cell-cell communications in order for tumour cells to successfully colonise distant host organs. Citation Format: Jean McBryan, Ailis Fagan, Damian McCartan, Jarlath C Bolger, Fiona T Bane, Christopher Byrne, Marie McIlroy, Arnold D Hill, Leonie S Young. Transcriptomic profiling of patient sequential tumours provides cutting edge view of global metastatic expression changes following endocrine therapy [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P2-05-13.

Abstract P3-06-01: Nuclear β-catenin negativity predicts for late relapse in ER+, tamoxifen-treated breast cancer

Abstract Background: The annual recurrence rates of post-menopausal ER-positive breast cancers persist beyond the first 5 years of diagnosis and treatment and the mortality rates in this period are higher versus ER-negative cancers. Extended endocrine therapy past 5 years has been shown to be of benefit but is associated with increased toxicity and cost thus the ability to predict patients who are at highest risk of late relapse would be of significant benefit in clinical decision making in this context. β-catenin is an intracellular protein that undergoes Wnt-mediated nuclear translocation where it transactivates genes implicated in tumour development and progression. We have also previously reported that β-catenin can play a role in aggressive resistance that accompanies prolonged endocrine treatment in vitro. In this study, we thus investigated whether β-catenin expression in ER+ breast cancer was predictive of recurrence beyond 5 years in an analysis of three separate endocrine-treated cohorts. Methods: Associations between β-catenin gene expression and relapse free survival (RFS) were performed using the online KMplotter tool. Immunostaining for total β-catenin was performed on tissue samples from 3 ER+ primary breast cancer series with long-term follow-up data: Nottingham 2000 (n=384, tamoxifen-only); ABC (n=570; tamoxifen only); transATAC (n=743; tamoxifen or ansatrozole). The association between subcellular (nuclear or cytoplasmic) β-catenin expression and RFS was determined in (i) the entire cohort, (ii) the first 5 years of tamoxifen treatment versus post-5 years. Results: KMplotter analysis of β-catenin in ER+, tamoxifen-treated patient samples (n=665) revealed a significant relationship with improved RFS in the post-five year cohort [HR: 0.48 (0.34-0.68); p=0.000019) versus the first 5 years [HR: 0.91 (0.61-1.36; p=0.64)]. No significant association was observed in untreated ER+ patients. In the Nottingham series, nuclear β-catenin positivity was significantly associated with improved survival in the 20-year follow-up for tamoxifen-treated patients (p=0.047) but not in the first five years (p=0.239). Cytoplasmic β-catenin did not associate with survival. Further analysis in the ABC series revealed an association between nuclear β-catenin positivity and improved survival for tamoxifen-treated patients in the entire cohort (HR: 0.52 (0.18, 0.55); p=0.00005). However, nuclear β-catenin was more strongly associated with improved outcome in the post 5-year treatment group (HR: 0.30 (0.14, 0.66); p=0.01) versus the first five years of treatment (HR: 0.33 (0.15,0.73); p=0.06). No significant associations were seen with cytoplasmic β-catenin. The transATAC trial material again revealed an association between presence of nuclear β-catenin and reduced distant recurrence in the post 5-years endocrine-treated cohort (HR: 0.54 (0.33, 0.91): x2=5.52, p=0.018) versus years 1-5 (HR: 1.16 (0.67, 2.01); x2=0.29, p=0.59). Conclusions: This is the first study to demonstrate that nuclear β-catenin may represent a predictive biomarker for late relapse following tamoxifen treatment in ER+ breast cancer where, contrary to traditional hypotheses and pre-clinical data, its nuclear expression is strongly associated with good outcome post-five years of treatment. Citation Format: Stephen Hiscox, Chris Smith, Robert I Nicholson, Julia Gee, Adrian Harris, Judith Bliss, Eleftheria Kalaizak, Ivana Sestak, Mitch Dowsett, Jack Cuzick, Ian Ellis, Peter Barrett-Lee. Nuclear β-catenin negativity predicts for late relapse in ER+, tamoxifen-treated breast cancer [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-06-01.

Abstract P1-03-05: Genetic variant in the OPG gene is associated with aromatase inhibitor-related musculoskeletal toxicity in breast cancer patients

Abstract Background Aromatase inhibitor (AI) therapy is associated with musculoskeletal toxicity, which adversely affects quality of life and therapy adherence. Our objective was to evaluate genetic predictors for endocrine therapy-related musculoskeletal pain in a prospective cohort study, which was previously published. Patients & methods 254 early breast cancer patients starting AI (n=159) or tamoxifen therapy (n=95) were included in this genetic biomarker study. Musculoskeletal symptoms were assessed at baseline and at 3, 6 and 12 months thereafter. AI-induced musculoskeletal pain was defined as an increase in arthralgia or myalgia relative to baseline. Single nucleotide polymorphisms (SNP) in candidate genes involved in estrogen signaling or reported to have associations with AI-related musculoskeletal pain or estrogen levels were selected. Of the 40 selected SNPs, 7 failed genotyping and were excluded from further analysis. Results Twenty SNPs exhibited a minor allele frequency <0.05 and were omitted from the final analysis. Overall, 13 SNPs in CYP19, CYP17, osteoprotegerin (OPG) and estrogen receptor 1 were included. 74% of AI-users reported new or worsened musculoskeletal pain compared to 33% of tamoxifen-treated patients. The OPG SNP rs2073618 was significantly associated with AI-induced musculoskeletal toxicity after correction for multiple testing (p=0.039). None of the other SNPs showed significant associations, both in AI and tamoxifen-treated patients. Conclusion The SNP rs2073618 in OPG predicts the risk of musculoskeletal symptoms with AI therapy, which has not been reported so far. Validation of this finding and further functional studies are needed. Citation Format: Anneleen Lintermans, Kathleen Van Asten, Annouschka Laenen, Thomas Van Brussel, Johan Verhaeghe, Dirk Vanderschueren, Diether Lambrechts, Patrick Neven. Genetic variant in the OPG gene is associated with aromatase inhibitor-related musculoskeletal toxicity in breast cancer patients [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P1-03-05.

Abstract P3-05-20: ESRP1 adds sp(l)ice to endocrine resistance

Abstract Introduction: De novo or acquired resistance to endocrine therapy limits its utility in a significant number of estrogen receptor (ER) breast cancers. An increasing number of molecular assays predict the likelihood of distant recurrence in tamoxifen-treated patients with node-negative, ER+ breast cancer. However, these do not provide the mechanistic basis for endocrine resistance. It is crucial to identify novel targets and improve the success of endocrine therapies. We previously shown that epithelial splicing regulatory proteins 1 and 2 (ESRP1 and ESRP2), RNA binding proteins that promote splicing, are significantly elevated in cases with high Oncotype DX scores (innate resistance) and in ERα-positive cells with acquired tamoxifen resistance (MCF-7 LCC2 cells) and fulvestrant and tamoxifen resistance (MCF-7/LCC9 cells). The aim of this study was to investigate the ESRP1/ESRP2-regulated alternative splicing events leading to innate and acquired endocrine resistance. Methods: A combinatorial bioinformatics approach was employed to identify genes altered through alternative splicing by ESRP1/ESRP2 in endocrine resistance. Briefly, genes with ESRP1/ESRP2 motifs were screened in the human genome. This data was integrated with tamoxifen-treated datasets, and filtered using protein-protein interactions (PPI; BIOGRID) and clinical outcome (KM plotter. Potential target gene transcripts were further narrowed down using an algorithm based on motif binding location and the Cancer Genome Atlas (TCGA) breast cancer datasets with low and high ESRP1 cases. The alternative transcripts were further validated using splice variant specific-custom qRT-PCR in low and high RS Oncotype cases as surrogate for endocrine sensitivity and resistance. Results: Motif analysis in combination with tamoxifen-treated datasets identified 2212 differentially expressed genes. Further collective analysis of number of PPI (>50) and survival data from KM plotter (P<0.001) narrowed down 79 candidate genes that are associated with tamoxifen resistance. TCGA data confirmed presence of distinct transcripts (splicing variants) based on ESPR1 expression levels. Splice specific RT-PCR confirmed alternative splicing events involving cycle related genes such as AURKA, FZR1, and MDM2 in low ESRP1 (and low RS) and high ESRP1 (high RS) cases. Conclusion: ESRP1/ESRP2 by inducing alternative splicing play an important role in tamoxifen resistance and recurrence of ER+ breast cancer. Targeting alternative splicing may offer novel avenues for combating endocrine-resistance in breast cancer. Citation Format: Yesim Gokmen-Polar, Yaseswini Neelamraju, Chirayu Pankaj Goswami, Harikrishna Nakshatri, Sarath Chandra Janga, Sunil Badve. ESRP1 adds sp(l)ice to endocrine resistance [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-05-20.

Abstract P6-01-10: Prognostic significance of breast cancer index (BCI) in node-positive hormone receptor positive early breast cancer: NCIC CTG MA.14

Abstract Background: The continuous linear Breast Cancer Index (BCI) risk index combines the ratio of genes HOXB13 to IL17BR (H/I) and the molecular grade index (MGI) (Zhang et al, Clinical Cancer Research, 2013). The BCI signature was developed for node-negative breast cancer patients treated with tamoxifen. We examine here whether linear BCI is prognostic for node-positive hormone-receptor positive tamoxifen-treated patients. Methods: MA.14 randomly assigned 667 hormone positive (HR+), postmenopausal women to 5 years of tamoxifen (TAM) +/- 2 years of octreotide LAR (TAM-OCT). A representative subgroup of 299 patients underwent gene expression profiling by RT-PCR for linear BCI. We performed exploratory analyses restricted to node positive patients. The primary objective was to assess the prognostic effect of BCI on relapse-free survival (RFS). RFS was defined as the time from randomization to the time of recurrence of the primary disease alone, including local and ipsilateral nodal recurrence and metastatic disease, and censoring at longest follow-up or death from another cause. With a median 9.8 years follow-up, the association of BCI with RFS was assessed by multivariate Cox regression including treatment, stratification factors (other than nodal status), and baseline patient and tumor characteristics. Patients were defined to be low risk based on BCI if the adjusted Cox survival was >95%, where adjustment was by trial treatment, stratification factors, and baseline patient and tumor characteristics, including IGF-1, IGFBP-3, and C-peptide. Results: 292 of 299 patient samples passed internal analytical quality control; 116 node positive ER+ve patients had 34 (29.3%) relapses, with adjusted Cox survival at 9.6 years of 87.8%. Fifty-two of the 116 patients (45%) did not receive adjuvant chemotherapy, and experienced 11 (21%) RFS events. In the 116 patients, higher continuous BCI value was associated with shorter RFS (p=0.002): hazard ratio (HR) 1.49 (95% CI 1.16-1.91). Smaller pathologic T had significantly (p=0.03) better RFS HR=0.39, (95%CI 0.17-0.90). With MA.14 patient mean BCI of 5.09532, Cox survival at 4.1 years was 95.2%; 17/34 (50%) who recurred had failed by this time. Discussion: In this subgroup analysis, we found that BCI and tumor size were significant prognostic factors for node-positive hormone-receptor positive patients who were treated with tamoxifen. Citation Format: Dennis Sgroi, Paul Goss, Judy-Anne Chapman, Elizabeth Richardson, Shemeica Binns, Yi Zhang, Cathy Schnabel, Mark Erlander, Kathy Pritchard, Lei Han, Lois Sheperd, Michael Pollack. Prognostic significance of breast cancer index (BCI) in node-positive hormone receptor positive early breast cancer: NCIC CTG MA.14 [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P6-01-10.

Caffeine and Caffeic Acid Inhibit Growth and Modify Estrogen Receptor and Insulin-like Growth Factor I Receptor Levels in Human Breast Cancer.

Epidemiologic studies indicate that dietary factors, such as coffee, may influence breast cancer and modulate hormone receptor status. The purpose of this translational study was to investigate how coffee may affect breast cancer growth in relation to estrogen receptor-α (ER) status. The influence of coffee consumption on patient and tumor characteristics and disease-free survival was assessed in a population-based cohort of 1,090 patients with invasive primary breast cancer in Sweden. Cellular and molecular effects by the coffee constituents caffeine and caffeic acid were evaluated in ER(+) (MCF-7) and ER(-) (MDA-MB-231) breast cancer cells. Moderate (2-4 cups/day) to high (≥5 cups/day) coffee intake was associated with smaller invasive primary tumors (Ptrend = 0.013) and lower proportion of ER(+) tumors (Ptrend = 0.018), compared with patients with low consumption (≤1 cup/day). Moderate to high consumption was associated with lower risk for breast cancer events in tamoxifen-treated patients with ER(+) tumors (adjusted HR, 0.51; 95% confidence interval, 0.26-0.97). Caffeine and caffeic acid suppressed the growth of ER(+) (P ≤ 0.01) and ER(-) (P ≤ 0.03) cells. Caffeine significantly reduced ER and cyclin D1 abundance in ER(+) cells. Caffeine also reduced the insulin-like growth factor-I receptor (IGFIR) and pAkt levels in both ER(+) and ER(-) cells. Together, these effects resulted in impaired cell-cycle progression and enhanced cell death. The clinical and experimental findings demonstrate various anticancer properties of caffeine and caffeic acid against both ER(+) and ER(-) breast cancer that may sensitize tumor cells to tamoxifen and reduce breast cancer growth.

Highlights of This Issue

Breast cancer remains the leading cause of cancer-related death among women worldwide. Modifiable dietary components, such as coffee, may influence breast cancer and treatment responses. In a translational study, Rosendahl and colleagues uncovered associations between coffee consumption and tumor characteristics in breast cancer patients, as well as longer disease-free survival among tamoxifen-treated patients. Mechanistically, this was linked to altered estrogen- and insulin-like growth factor I receptor levels and downstream signaling, resulting in impaired cell-cycle progression and enhanced cell death. An improved understanding of how coffee can influence breast cancer and treatment response may yield more evidence-based lifestyle recommendations during treatment.It has long been advocated that normalization of peritumoral vessels through vascular endothelial growth factor (VEGF) inhibition may help to improve cytotoxic drug delivery to tumors. The best timing of giving VEGF inhibitor, however, remains unknown. In this clinical trial, Lu and colleagues tackled this issue by administrating bevacizumab one day before the combination of etoposide and cisplatin (BEEP regimen) to breast cancer patients whose metastatic brain tumors were refractory to whole brain radiotherapy. The BEEP regimen showed a central nervous system response rate of 77.1% by volumetric criteria and was associated with manageable toxicities.The pediatric tumor neuroblastoma presents with extraordinarily heterogeneous clinical courses, ranging from spontaneous regression to fatal disease progression. Accurate assessment of each individual patient's risk is therefore essential to appropriately tailor therapeutic intensities. To improve treatment stratification, Oberthuer and colleagues determined gene expression profiles of 709 primary neuroblastomas by microarrays and developed a gene expression-based classifier that predicts patient outcome with unprecedented precision. They developed a novel risk assessment system that combines the classifier with established markers, which may avoid over- and under-treatment of future neuroblastoma patients.Analyzing multiple tumor regions, Friemel and colleagues demonstrate intratumor heterogeneity in the majority of treatment-naïve hepatocellular carcinomas (HCC). Intratumor heterogeneity was not restricted to morphological features, immunohistochemical markers, or mutational aberrations of critical driver genes, CTNNB1 and TP53, but hierarchically linked between all three. The emergence of intratumor heterogeneity highlights the challenges associated with molecular classification of HCC and the development of targeted therapies because distinct HCC subpopulations may foster tumor adaptations resulting in therapy resistance or evasion.

Reprogramming of the ERRα and ERα target gene landscape triggers tamoxifen resistance in breast cancer.

Endocrine treatment regimens for breast cancer that target the estrogen receptor-α (ERα) are effective, but acquired resistance remains a limiting drawback. One mechanism of acquired resistance that has been hypothesized is functional substitution of the orphan receptor estrogen-related receptor-α (ERRα) for ERα. To examine this hypothesis, we analyzed ERRα and ERα in recurrent tamoxifen-resistant breast tumors and conducted a genome-wide target gene profiling analysis of MCF-7 breast cancer cell populations that were sensitive or resistant to tamoxifen treatment. This analysis uncovered a global redirection in the target genes controlled by ERα, ERRα, and their coactivator AIB1, defining a novel set of target genes in tamoxifen-resistant cells. Beyond differences in the ERα and ERRα target gene repertoires, both factors were engaged in similar pathobiologic processes relevant to acquired resistance. Functional analyses confirmed a requirement for ERRα in tamoxifen- and fulvestrant-resistant MCF-7 cells, with pharmacologic inhibition of ERRα sufficient to partly restore sensitivity to antiestrogens. In clinical specimens (n = 1041), increased expression of ERRα was associated with enhanced proliferation and aggressive disease parameters, including increased levels of p53 in ERα-positive cases. In addition, increased ERRα expression was linked to reduced overall survival in independent tamoxifen-treated patient cohorts. Taken together, our results suggest that ERα and ERRα cooperate to promote endocrine resistance, and they provide a rationale for the exploration of ERRα as a candidate drug target to treat endocrine-resistant breast cancer.

Florid Endocervical Microglandular Hyperplasia in Association with Tamoxifen Treatment

Background: Although tamoxifen has well documented oestrogen-like effects in the endometrium and ectocervical squamous epithelium, its effects on the endocervix are poorly documented. Case report: We report a case of an 88 year-old woman who presented with postmenopausal bleeding after receiving tamoxifen for 16 months following a diagnosis of Grade 2 invasive and in situ carcinoma of breast. Endometrial biopsy was performed revealing either atypical hyperplasia or adenocarcinoma. She underwent total abdominal hysterectomy, bilateral salpingo-oophorectomy and lymph gland sampling. Histopathological examination of the uterus showed an endometrioid carcinoma (FIGO Grade 2) at the endometrium. In addition, there was extensive florid endocervical microglandular hyperplasia. Conclusion: Endocervical microglandular hyperplasia is usually seen in woman of reproductive age, particularly in association with pregnancy and contraceptive use. It is unusual in postmenopausal woman and its presence in this 88 year-old tamoxifentreated patient implicates tamoxifen as a course of endocervical microglandular hyperplasia in this instance.

The potential role of estrogen receptor β2 in breast cancer

Endocrine therapy is provided to all patients with estrogen receptor (ER)-positive breast cancer, but only a subset of them derives clinical benefit. The discovery of ERβ and its five isoforms added another layer of complexity in the regulation of estrogen activity in breast cancer cells. Two large retrospective studies showed conflicting results with regard to the prognostic value of the different ERβ isoforms in patients treated with tamoxifen in an adjuvant setting. This study tested the hypothesis that ERβ1 and, or ERβ2 are correlated with clinical outcome. We identified patients with breast cancer who had undergone surgery at Bucheon St. Mary's Hospital, the Catholic University of Korea, between January 2004 and March 2006. We evaluated 101 consecutive cases for ERβ1 and ERβ2 expression using immunohistochemical staining and obtained other clinicopathology by reviewing medical records. ERβ1 was expressed in 81.2% (79 of 97) and ERβ2 was expressed in 50.5% (51 of 101) of primary breast cancer tissues. Disease-free survival (DFS) and overall survival (OS) of patients with cancers expressing ERβ2 was significantly worse. Moreover, in subgroup analysis according to the tamoxifen treatment, ERβ2 expression was significantly associated with shorter DFS of tamoxifen-treated patients. This study indicates that breast cancer with ERβ2 expression was associated with worse DFS and OS, especially in tamoxifen treated patients. Our results suggest a role for ERβ2 as an independent prognostic marker and might serve as a new therapeutic target.

ERβ1: characterization, prognosis, and evaluation of treatment strategies in ERα-positive and -negative breast cancer.

BackgroundThe role and clinical value of ERβ1 expression is controversial and recent data demonstrates that many ERβ antibodies are insensitive and/or non-specific. Therefore, we sought to comprehensively characterize ERβ1 expression across all sub-types of breast cancer using a validated antibody and determine the roles of this receptor in mediating response to multiple forms of endocrine therapy both in the presence and absence of ERα expression.MethodsNuclear and cytoplasmic expression patterns of ERβ1 were analyzed in three patient cohorts, including a retrospective analysis of a prospective adjuvant tamoxifen study and a triple negative breast cancer cohort. To investigate the utility of therapeutically targeting ERβ1, we generated multiple ERβ1 expressing cell model systems and determined their proliferative responses following anti-estrogenic or ERβ-specific agonist exposure.ResultsNuclear ERβ1 was shown to be expressed across all major sub-types of breast cancer, including 25% of triple negative breast cancers and 33% of ER-positive tumors, and was associated with significantly improved outcomes in ERα-positive tamoxifen-treated patients. In agreement with these observations, ERβ1 expression sensitized ERα-positive breast cancer cells to the anti-cancer effects of selective estrogen receptor modulators (SERMs). However, in the absence of ERα expression, ERβ-specific agonists potently inhibited cell proliferation rates while anti-estrogenic therapies were ineffective.ConclusionsUsing a validated antibody, we have confirmed that nuclear ERβ1 expression is commonly present in breast cancer and is prognostic in tamoxifen-treated patients. Using multiple breast cancer cell lines, ERβ appears to be a novel therapeutic target. However, the efficacy of SERMs and ERβ-specific agonists differ as a function of ERα expression.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-749) contains supplementary material, which is available to authorized users.

Protein kinase D1 regulates ERα-positive breast cancer cell growth response to 17β-estradiol and contributes to poor prognosis in patients.

About 70% of human breast cancers express and are dependent for growth on estrogen receptor α (ERα), and therefore are sensitive to antiestrogen therapies. However, progression to an advanced, more aggressive phenotype is associated with acquisition of resistance to antiestrogens and/or invasive potential. In this study, we highlight the role of the serine/threonine-protein kinase D1 (PKD1) in ERα-positive breast cancers. Growth of ERα-positive MCF-7 and MDA-MB-415 human breast cancer cells was assayed in adherent or anchorage-independent conditions in cells overexpressing or depleted for PKD1. PKD1 induces cell growth through both an ERα-dependent manner, by increasing ERα expression and cell sensitivity to 17β-estradiol, and an ERα-independent manner, by reducing cell dependence to estrogens and conferring partial resistance to antiestrogen ICI 182,780. PKD1 knockdown in MDA-MB-415 cells strongly reduced estrogen-dependent and independent invasion. Quantification of PKD1 mRNA levels in 38 cancerous and non-cancerous breast cell lines and in 152 ERα-positive breast tumours from patients treated with adjuvant tamoxifen showed an association between PKD1 and ERα expression in 76.3% (29/38) of the breast cell lines tested and a strong correlation between PKD1 expression and invasiveness (P < 0.0001). In tamoxifen-treated patients, tumours with high PKD1 mRNA levels (n = 77, 50.66%) were significantly associated with less metastasis-free survival than tumours with low PKD1 mRNA expression (n = 75, 49.34%; P = 0.031). Moreover, PKD1 mRNA levels are strongly positively associated with EGFR and vimentin levels (P < 0.0000001). Thus, our study defines PKD1 as a novel attractive prognostic factor and a potential therapeutic target in breast cancer.

Arthralgia induced by endocrine treatment for breast cancer: A prospective study of serum levels of insulin like growth factor-I, its binding protein and oestrogens

BackgroundAromatase inhibitors (AIs) frequently induce or enhance musculoskeletal problems (AI-induced musculoskeletal syndrome (AIMSS)) which sometimes are debilitating. Apart from low oestrogen levels, underlying mechanisms are unknown and likely multiple. We previously hypothesised a role for the growth hormone/insulin like growth factor-I (IGF-I) axis. Here, we report the effect of tamoxifen and AI on IGF-I, IGF binding protein-3 (IGFBP-3) and oestrogen levels from a prospective study. Materials and methodsPostmenopausal women with an early breast cancer scheduled to start adjuvant endocrine therapy with an AI or tamoxifen were recruited. A rheumatologic questionnaire was completed and serum was collected for assessment of IGF-I, IGFBP-3 and oestrogen levels. Re-evaluation was done after 3, 6 and 12months of therapy. Results84 patients started on tamoxifen (n=42) or an AI (n=42). 66% of the latter group experienced worsening of pre-existing or de novo complaints in joint and/or muscle, compared to 29% of tamoxifen-treated patients. AI therapy resulted in elevated IGF-I levels with a statistically significant increase at 6months (p=0.0088), whereas tamoxifen users were characterised by a decrease in IGF-I levels at all follow-up times (p<0.0004). No effect on IGFBP-3 was seen in the latter group. AI-users, however, showed decreased IGFBP-3 levels at 12months (p=0.0467). AIMSS was characterised by a decrease in IGFBP-3 levels (p=0.0007) and a trend towards increased IGF-I/IGFBP-3 ratio (p=0.0710). ConclusionThese findings provide preliminary evidence that AI-induced musculoskeletal symptoms are associated with changes in the growth hormone (GH)/IGF-I axis.

Abstract 1887: Global gene expression analysis of tamoxifen-resistant breast cancer cell lines identified a gene signature predictive of clinical outcome of breast cancer patients treated with endocrine therapy

Abstract Objective: Altered gene expression associated with endocrine resistance in breast cancer cell lines may have potential prognostic/predictive values in patients with ER+ breast cancer receiving endocrine therapy. In this study, we investigated mRNA expression profiles associated with the tamoxifen-resistant phenotype using a panel of isogenic MCF-7-based breast cancer cell lines to elucidate the endocrine resistance mechanism and assess whether the pattern of gene expression predicted clinical outcome in breast cancer patients treated with endocrine therapy. Method: Gene array was performed on a panel of clinically-relevant tamoxifen-resistant breast cancer cell lines (TamR) and their tamoxifen-sensitive parental cell line. Genes that showed significant alteration in expression (FDR 0.01) in TamRs vs parental cell lines were evaluated for their potential to stratify tamoxifen-treated breast cancer patients based on outcome using published gene expression data sets. Results: Using a testing set consisting of microarray data from 163 ER+ primary breast cancer patients treated with tamoxifen we identified 20 genes among those that showed significant altered expression in TamR that could stratify patients according to clinical outcome. . The potential of this 20-gene signature to stratify patients was further confirmed in 2 independent data sets consisting of ER+ tamoxifen-treated breast cancer patients with a sensitivity and specificity 53% and 79% respectively in the first cohort (N=153), and sensitivity and specificity of 37% and 91% respectively in the second cohort (N=201). Evaluation of relapse-free survival of tamoxifen-treated patients classified by the signature as having bad or good outcomes showed significant differences in time-to-relapse, with median disease-free survival times of 5.1 vs 8.9 years in the first cohort (N=153, p=0.03) and 4.2 vs 7.7 years in the second (N=201, p=0.006). Conclusions: The expression pattern of the 20-gene signature identified in this study appears to have a strong potential as a biomarker of clinical outcome of endocrine-treated breast cancer patients. Citation Format: Daniel Elias Roro, Henriette Vever, Anne Lykkesfeldt, Henrik Ditzel. Global gene expression analysis of tamoxifen-resistant breast cancer cell lines identified a gene signature predictive of clinical outcome of breast cancer patients treated with endocrine therapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1887. doi:10.1158/1538-7445.AM2014-1887

Abstract 3295: S6 kinase signaling in prognosis and tamoxifen response in two randomized breast cancer cohorts

Abstract Detecting signals in the mammalian target of rapamycin (mTOR), and the estrogen receptor (ER) pathways for prediction of treatment response may be a future clinical tool in primary breast cancer. Here, we investigated the validity and value of the mTOR targets p70-S6 kinase (S6K) 1 and 2 as biomarkers for tamoxifen sensitivity in vitro and in two independent tamoxifen randomized postmenopausal breast cancer cohorts. In addition, the prognostic value of the S6Ks was evaluated. We found S6K1 to correlate with HER2 and cytoplasmic Akt activity, whereas S6K2 and phosphorylated S6K were closer connected with ER positivity, low proliferation and nucleic p-Akt. Treatment prediction and prognosis were evaluated by immunohistochemical staining. Nuclear accumulation of S6K1 was indicative of a reduced tamoxifen treatment effect, compared with a significant benefit from tamoxifen treatment in patients without tumor S6K1 nuclear accumulation. Patients with a combination of S6K1 nuclear accumulation and S6K2 cytoplasmic accumulation in the tumor cells had no tamoxifen benefit. Also, S6K1 and S6K2 activation, indicated by p-S6K-t389 expression, was associated with low benefit from tamoxifen compared with untreated patients. In addition, high protein expression of S6K1, independent of localization, predicted worse prognosis. This was not evident for variations in S6K2 or p-S6K-t389 expression. In conclusion, the mTOR targeted kinases S6K1 and S6K2 interfere with proliferation and response to tamoxifen. Monitoring their activity and intracellular localization may provide biomarkers for breast cancer treatment, allowing for identification of a group of patients less likely to benefit from tamoxifen and thus in need of an alternative or additional treatment. Citation Format: Josefine Bostner, Elin Karlsson, Cecilia Bivik, Gizeh Perez-Tenorio, Tommy Fornander, Olle Stål. S6 kinase signaling in prognosis and tamoxifen response in two randomized breast cancer cohorts. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3295. doi:10.1158/1538-7445.AM2014-3295

Abstract 562: Impact of ERα expression status and ESR1 genetic variation on progression in tamoxifen-treated breast cancer patients

Abstract Background. Resistance to tamoxifen remains a major clinical challenge in the treatment of breast cancer. The aim of this study was to evaluate the influence of the intensity and the pattern of estrogen receptor α (ERα) expression and common single nucleotide polymorphisms (SNP) in the estrogen receptor 1 gene (ESR1, rs2228480:2014G&amp;gt;A; rs2077647:+30T&amp;gt;C) on disease progression in breast cancer patients who received adjuvant tamoxifen. Material and methods. Formalin-fixed paraffin-embedded tumor tissue samples were retrospectively collected from 61 patients, who had distant metastasis versus distant metastasis-free breast cancers (n = 37 versus n = 24, respectively, with a 9-year follow-up). Protein expression of ER, PR and HER2 were assessed by immunohistochemistry. Real-time polymerase chain reaction analysis was used to determine the ESR1 SNPs. Results. We found that the heterogeneous distribution of ERα was statistically significant related with a low expression level and intensity of ERα in tumor tissue samples (p=0.002 and p=0.0002, respectively). In addition, the level and intensity of ERα expression was lower in patients with distant metastasis compared to distant metastasis-free cases, but these differences were not significant (P&amp;gt;0.05). Nonetheless, the ESR1 2014A allele was associated with a heterogeneous distribution of ERα expression (p=0.02). We showed the higher risk of distant metastasis development for the patients carrying the ESR1 2014A mutant allele compared with women with ESR1 2014G variant (p=0.04). Conclusion. These data suggest that the expression level and intensity of ERα with ESR1 2014G&amp;gt;A genetic variation could be a useful molecular markers for predicting tamoxifen treatment prognosis in breast cancer patients. Citation Format: Nataliya Babyshkina, Sergey Vtorushin, Stanislav Patalyak, Tatyana Dronova, Elena Slonimskaya, Nadejda Cherdyntseva. Impact of ERα expression status and ESR1 genetic variation on progression in tamoxifen-treated breast cancer patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 562. doi:10.1158/1538-7445.AM2014-562

Abstract 403: Expression of genes spanning a breast cancer susceptibility locus on 6q25.1 is modulated by epigenetic modification

Abstract Single nucleotide polymorphisms within the region immediately upstream of the gene encoding ERα (ESR1) on 6q25.1, have been found to be associated with increased risk of breast cancer1,2. Three genes within this region, CCDC170, RMND1 and C6ORF211 are co-expressed with ESR1 in oestrogen receptor positive (ER+ve) breast cancers and CCDC170 expression is inversely associated with proliferation and recurrence free survival in a tamoxifen-treated patient cohort3. Epigenetic modification represents one potential mechanism through which expression of co-localised genes could be modulated. We investigated methylation and chromatin structure in the 6q25.1 using methylation sensitive sequencing, chromatin immunoprecipitation (ChIP) and chromatin confirmation capture (3C). Gene expression of ESR1, the three genes and alternative ESR1 mRNA isoforms were analysed in vitro using a panel of breast cancer cell lines. Long-term oestrogen deprived (LTED) cell lines were also generated and used to model the effects of anti-oestrogen therapies used in the treatment of ER positive breast cancers. DNA methylation at the ESR1/CCDC170 locus, gene expression of the four transcripts and alternative ESR1 mRNA isoforms differed greatly between the breast cancer cell lines. ER negative breast cancer cell lines were similar to the ‘non-tumorigenic’ MCF10A cell line while ER positive cell lines were hypomethylated and showed higher expression relative to MCF10A. We also discovered two putative regulatory binding regions which were differentially methylated that correlated with gene expression in our cell lines; one negatively and one positively. Analysis of ER positive tumors found that oestrogen receptor positive tumors were hypomethylated relative to normal tissue. In tumors, two differentially methylated regions were shown to have a strong negative correlation with gene expression of ESR1 and the three genes. These gave correlation coefficients of -0.577, -0.593, -0.600 and -0.454 for ESR1, CCDC170, C6ORF211 and RMND1, respectively, using Pearson correlation analysis and correction for multiple testing. The differentially methylated regions in tumors coincided with binding sites for ERα and Cohesin and long range chromatin interaction sites involving RNA polymerase II. Therefore, reduced DNA methylation seen in the ER positive tumors could facilitate enhanced chromatin interactions and transcription leading to the co-expression of the transcriptional hub. These alterations seen in the 6q25.1 locus, both in vitro and in our dataset, distinguish ER positive tumors and may contribute to resistance to anti-oestrogen therapies.

The use of Chinese herbal products and its influence on tamoxifen induced endometrial cancer risk among female breast cancer patients: A population-based study

Ethnopharmacological relevanceThe increased practice of traditional Chinese medicine (TCM) worldwide has raised concerns regarding herb–drug interactions. The purpose of our study was to analyze the use of Chinese herbal products (CHPs) and to estimate the influence of the use of CHP on tamoxifen induced endometrial cancer risk among female breast cancer patients in Taiwan. MethodsAll patients newly diagnosed with invasive breast cancer receiving tamoxifen treatment from January 1, 1998 to December 31, 2008 were selected from the National Health Insurance Research Database. The usage, frequency of service, and CHPs prescribed among the 20,466 tamoxifen-treated female breast cancer patients were analyzed. The logistic regression method was employed to estimate the odds ratios (ORs) for utilization of CHPs. Cox proportional hazard regression was performed to calculate the hazard ratios (HRs) for subsequent endometrial cancer for CHP non-users and CHP users among female breast cancer patients who had undergone tamoxifen treatment. ResultsMore than half of the subjects had ever used a CHP. Jia-Wei-Xiao-Yao-San (Augmented Rambling Powder) and Shu-Jing-Huo-Xue-Tang (Channel-Coursing Blood-Quickening Decoction) were the two most commonly used CHPs. The HR for the development of endometrial cancer among CHP users was 0.50-fold (95% CI=0.38–0.64) compared to that of CHP non-users. ConclusionMore than half of the study subjects had ever used a CHP. Jia-Wei-Xiao-Yao-San was the most commonly used CHP. Among female breast cancer patients who had undergone tamoxifen therapy, CHP consumption decreased the risk of subsequent endometrial cancer. Exploring potential Chinese herb–tamoxifen interactions and integrating both healthcare approaches are beneficial to the overall health outcomes of tamoxifen-treated female breast cancer patients.