The Journal of Steroid Biochemistry and Molecular Biology
Gene network signaling in hormone responsiveness modifies apoptosis and autophagy in breast cancer cells☆
Introduction
Over 40,000 American women die of breast cancer each year [1]; incidence is broadly similar across the European Union when considered as a percentage of the population. In 2008, over 178,000 women will be diagnosed with invasive breast cancer in the U.S., almost 70% of which will be estrogen receptor-α positive (ER+; HUGO Gene Symbol = ESR1) [2], [3]. The percentage of ER+ sporadic breast cancers increases linearly with age but even in premenopausal cases the proportion is high; 62% at age ≤35 and 72% by age 49 [2], [3], [4]. Data from randomized trials and meta-analyses clearly show that all breast cancer patients derive a statistically significant survival benefit from adjuvant chemotherapy, and that all hormone receptor positive breast cancer patients benefit from adjuvant endocrine therapy [5], [6], [7], [8], [9]. For postmenopausal women, the benefit from adjuvant Tamoxifen (TAM) is comparable to that seen for cytotoxic chemotherapy. While 5 years of adjuvant TAM produces a 26% proportional reduction in mortality [8], many ER+ tumors eventually recur [10]. Since advanced ER+ breast cancer largely remains an incurable disease, resistance to endocrine therapies is a significant clinical problem.
Endocrine therapy is administered as an antiestrogen (AE) like Tamoxifen (TAM) or Fulvestrant (FAS; Faslodex; ICI 182,780), or as an aromatase inhibitor (AI) such as Letrozole or Exemestane. It is less toxic and potentially more effective therapy in the management of hormone-dependent breast cancers. Antiestrogens, and TAM in particular, have been the “gold standard” first line endocrine therapy for over 30 years [11], clinical experience with this drug likely exceeding over 15 million patient years [10]. TAM increases both disease free and overall survival from early stage breast cancer, and it also reduces the incidence of invasive and noninvasive breast cancer in high-risk women [8], [9]. Raloxifene, another antiestrogen, is effective in reducing the rate of postmenopausal bone loss from osteoporosis as well as the rate of invasive breast cancer [12]. Newer antiestrogens such as FAS show significant activity relative to TAM and some AIs [13], [14]. Third generation AIs are now widely accepted as viable alternatives to AEs for first line endocrine therapy in postmenopausal women with metastatic disease; overall response rates are generally greater for AIs [15]. Importantly, Tamoxifen is the only single agent with demonstrated efficacy in both premenopausal and postmenopausal women with invasive breast cancer. Other AEs and all of the AIs require the complete cessation of ovarian function.
Of current interest is identification of the optimum choice and scheduling of AEs and AIs. Evidence clearly shows improvements in disease free survival for combined adjuvant therapy (an AI and an AE usually given sequentially) over single agent TAM [16], [17], [18], [19], [20]. However, the ability of AIs to induce a significant improvement in overall survival compared with 5 years of TAM alone is uncertain [15]. In terms of metastatic disease, recent data imply that response rates with an AI are either equivalent with or higher than with TAM [21], [22]. Given the increasing number of endocrine treatment options, there is a clear need to optimize the selection and scheduling of agents for both early stage and advanced disease. Whichever way these controversies are eventually resolved, it is clear that both AIs and AEs will remain as key modalities in the management of ER+ breast cancers. Unfortunately, the inability of endocrine therapies to cure many women with ER+ disease will also remain.
Several resistance phenotypes are evident from both experimental models and clinical observations. The two primary receptor phenotypes are ER+ and ER−. These receptor-based phenotypes have been further stratified by addition of the estrogen-regulated receptor for progesterone (PGR; HUGO Gene Symbol = PGR). The degree of treatment benefit from endocrine therapy varies according to receptor phenotype. For example, approximately 75% of ER+/PGR+, 33% of ER+/PGR−, and 45% of ER−/PGR+ cases of metastatic breast cancer respond to TAM [10]. Endocrine responses in truly ER− tumors are probably relatively rare and of uncertain relevance, as they most likely reflect incorrect assessments of what may be very low ER and/or PGR expression values. Data from the Early Breast Cancer Trialists’ Collaborative Group meta-analyses show that TAM therapy generates a non-significant 6% reduction in the 10-year risk of recurrence. A non-significant increase in the risk of death from any cause in patients with ER− breast cancer also was reported [8], [9]. The real value of PGR, which is the only modification to this clinical prediction scheme for directing endocrine therapy to become routine in over 30 years (the value of directing endocrine therapy based on HER2 is still controversial), is largely limited to ER− tumors. It is general practice in the United States to treat all ER+ and/or PR+ invasive breast tumors with endocrine therapy. However, it remains impossible to predict whether an individual patient will receive benefit from treatment and the magnitude or duration of any benefit. Better predictors of each individual patient's endocrine responsiveness are clearly needed.
Several pharmacological phenotypes have been identified in experimental models of either human breast cancer cells growing in vitro or of xenografts in immune-deficient rodents [10]. These phenotypes include (i) estrogen-independent (which appears equivalent to AI resistance but is not so for antiestrogen resistance [23]—some breast cancers can become resistant to an AE but still respond to an AI and vice versa); (ii) estrogen-inhibited (recently identified in MCF-7 models [24]); (iii) TAM-stimulated (identified first in MCF-7 xenografts [25], [26]); TAM-unresponsive but FAS sensitive [27] (identified first in MCF-7 models and subsequently observed in clinical trials [13]); TAM and FAS crossresistant [28] (perhaps this is truly antiestrogen crossresistant and it is seen both clinically in patients and experimentally in MCF-7 models [13], [29]). Other variations on these phenotypes likely occur but are beyond the scope of our discussion.
Obtaining direct clinical evidence for the prevalence of each of the pharmacological resistance phenotypes is challenging. We have previously noted the utility of applying clinical responses to TAM withdrawal in metastatic breast cancer as one means to define, at least in broad terms, the likely relevance of a series of pharmacological phenotypes [29]. This approach is somewhat limited, as the number of cases across all studies is modest (n = 241). Furthermore, TAM withdrawal responses cannot readily distinguish between TAM-stimulation and estrogen-inhibition because each should predict for a clinical benefit. The latter would induce a benefit because many breast cancers contain significant concentrations of 17β-estradiol, independent of both menopausal and ER/PGR status [10], sufficient to produce the estrogen-inhibited phenotype [24]. Indeed, the superiority of AIs over TAM in inducing clinical response strongly implies that over 75% of ER+/PGR+, at least 50% of all ER+ breast cancers irrespective of PGR expression, and 45% or more of ER−/PGR+ breast tumors are probably driven by adequate access to estrogen.
In our prior assessment, almost 9% of patients received an overall clinical response to TAM withdrawal (partial responses + complete responses). When disease stabilizations were included we estimated that less than 20% of patients received clinical benefit [29], suggesting that the sum of TAM-stimulated plus estrogen-inhibited clinical phenotypes may not account for the majority of resistant phenotypes in women. Of course, given the number of ER+ breast cancers arising every year, these phenotypes are relevant to a notable number of women. The major response to TAM withdrawal was clinically detectable disease progression – greater than 80% of cases – strongly implicating unresponsiveness as the primary clinical resistance mechanism to TAM. Whether these breast cancers are fully crossresistant to all endocrine therapies, or retain sensitivity to AIs, cannot be determined from this simple analysis.
Nomura et al. [30] took a different approach and assessed the responsiveness to estrogen and TAM in short-term primary cell cultures of n = 153 ER+ breast cancer biopsies. This approach allowed the authors to separate the various pharmacological phenotypes; approximately 7% of ER+ primary cultures were stimulated by TAM and almost 3% were inhibited by physiological concentrations of estradiol—notably close to our estimate of 9% for the sum of these two clinical phenotypes.
It is important here to separate responses to physiological estrogens from those produced by pharmacological estrogen therapy. High dose estrogen therapy was used prior to the advent of TAM. As with all endocrine therapies, approximately 30% of all breast cancers (receptor status was not available when most of these studies were done) responded [31], [32]. Side effects were unfavorable, probably explaining the switch to TAM that also induces responses in approximately 30% of all breast cancers (when receptor status is not considered). It is also likely that the mechanisms of action of pharmacological and physiological dose estrogens differ. Over 15 years ago, we were the first to show that pharmacological concentrations of both estradiol and TAM induce changes in the membrane fluidity of breast cancer cells and that this correlates with changes in cell growth [33]. It is unlikely that membrane fluidity changes are major contributors to the action, either prosurvival or prodeath, of physiological estrogen exposures but they likely do contribute to the prodeath effects of pharmacological exposures.
Section snippets
Cell fate in the context of endocrine responsiveness
Therapeutic strategies for breast cancer generally aim to alter the balance between cell death and cell survival such that cancer cells (but ideally not normal cells) die. However, endocrine therapies consistently also induce a notable growth arrest in sensitive tumors. The relative importance of growth arrest and cell death remains unclear. To explore this issue, we will first discuss the forms of cell death and then compare the potential for cell death and cell growth arrest to contribute to
Molecular signaling and resistance
The precise mechanisms of resistance to an AE and/or an AI remain unclear, reflecting an incomplete understanding of the signaling affecting cell proliferation, survival, and death and their hormonal regulation in breast cancer. We have previously reviewed the mechanisms of resistance to AEs and to estrogen deprivation elsewhere in some detail [10], [23], [29], so we focus here on the molecular signaling aspects of resistance and how these may be integrated and explored using emerging
Seed-gene model for cell signaling and the regulation of cell fate
While we continue to develop new methods for network modeling, we have yet to report our modeling approaches to our own expanding data sets. Hence, we will here describe our initial studies on the use of seed genes and experimental data to construct a simple wiring-diagram of our initial seed-gene network. The inability to induce signaling to irreversible cell death is a central component of drug resistance [94]. Thus, we propose that cells possess a common cell death/survival regulatory
References (160)
- et al.
Response to a specific antioestrogen (ICI 182,780) in tamoxifen-resistant breast cancer
Lancet
(1995) - et al.
Switching of postmenopausal women with endocrine-responsive early breast cancer to anastrozole after 2 years’ adjuvant tamoxifen: combined results of ABCSG trial 8 and ARNO 95 trial
Lancet
(2005) - et al.
Antiestrogens, aromatase inhibitors, and apoptosis in breast cancer
Vitam. Horm.
(2005) - et al.
Bcl-2 antiapoptotic proteins inhibit Beclin 1-dependent autophagy
Cell
(2005) - et al.
Death by design: apoptosis, necrosis and autophagy
Curr. Opin. Cell Biol.
(2004) - et al.
If not apoptosis, then what? Treatment-induced senescence and mitotic catastrophe in tumor cells
Drug Resist. Updat.
(2001) - et al.
Structural basis for sorting mechanism of p62 in selective autophagy
J. Biol. Chem.
(2008) - et al.
Effective tamoxifen therapy of breast cancer involves both antiproliferative and pro-apoptotic changes
Eur. J. Cancer
(2000) - et al.
Molecular and pharmacological aspects of antiestrogen resistance
J. Steroid Biochem. Mol. Biol.
(2001) - et al.
The A4396G polymorphism in interferon regulatory factor-1 is frequently expressed in breast cancer
Cancer Genet. Cytogenet.
(2007)
Cancer statistics, 2006
CA Cancer J. Clin.
Estrogen receptor status in BRCA1- and BRCA2-related breast cancer: the influence of age, grade, and histological type
Clin. Cancer Res.
Estrogen and progesterone receptor determinations in breast cancer. Technology, biology and clinical significance
Acta Oncol.
Re: Estrogen receptor status of primary breast cancer is predictive of estrogen receptor status of contralateral breast cancer
J. Natl. Cancer Inst.
Polychemotherapy for early breast cancer: an overview of randomised trials
Lancet
Sequential methotrexate and fluorouracil for the treatment of node-negative breast cancer patients with estrogen-receptor-negative tumors: eight year results from National Surgical Adjuvant Breast and Bowel Project (NSABP) B-13 and first report of findings from NSABP B-19 comparing methotrexate and fluorouracil with conventional cyclophosphamide, methotrexate, and fluorouracil
J. Clin. Oncol.
Survival advantage of adjuvant chemotherapy in high-risk node-negative breast cancer: ten-year analysis—an intergroup study
J. Clin. Oncol.
Tamoxifen for early breast cancer: an overview of the randomized trials
Lancet
Systemic treatment of early breast cancer by hormonal, cytotoxic, or immune therapy
Lancet
Cellular and molecular pharmacology of antiestrogen action and resistance
Pharmacol. Rev.
A new antioestrogenic agent in late breast cancer. An early clinical appraisal of ICI 46474
Br. J. Cancer
The effect of raloxifene on risk of breast cancer in postmenopausal women; results from the MORE randomized trial. Multiple Outcomes of Raloxifene Evaluation
J. Am. Med. Assoc.
Fulvestrant, formerly ICI 182, 780, is as effective as anastrozole in postmenopausal women with advanced breast cancer progressing after prior endocrine treatment
J. Clin. Oncol.
Second- and third-generation aromatase inhibitors as first-line endocrine therapy in postmenopausal metastatic breast cancer patients: a pooled analysis of the randomised trials
Br. J. Cancer
A comparison of letrozole and tamoxifen in postmenopausal women with early breast cancer
N. Engl. J. Med.
A randomized trial of exemestane after two to three years of tamoxifen therapy in postmenopausal women with primary breast cancer
N. Engl. J. Med.
Sequential tamoxifen and aminoglutethimide versus tamoxifen alone in the adjuvant treatment of postmenopausal breast cancer patients: results of an Italian cooperative study
J. Clin. Oncol.
Anastrozole alone or in combination with tamoxifen versus tamoxifen alone for adjuvant treatment of postmenopausal women with early-stage breast cancer: results of the ATAC (Arimidex, Tamoxifen Alone or in Combination) trial efficacy and safety update analyses
Cancer
Anastrozole is superior to tamoxifen as first-line therapy in hormone receptor positive advanced breast carcinoma
Cancer
Superior efficacy of letrozole versus tamoxifen as first-line therapy for postmenopausal women with advanced breast cancer: results of a phase iii study of the international letrozole breast cancer group
J. Clin. Oncol.
Cross resistance and molecular mechanisms in antiestrogen resistance
Endocr. Relat. Cancer
Paradoxical action of fulvestrant in estradiol-induced regression of tamoxifen-stimulated breast cancer
J. Natl. Cancer Inst.
Development of tamoxifen-stimulated growth of MCF-7 tumors in athymic mice after long-term antiestrogen administration
Cancer Res.
Human breast cancer in athymic nude mice: cytostatic effects of long-term antiestrogen therapy
Eur. J. Cancer Clin. Oncol.
MCF7/LCC2: A 4-hydroxytamoxifen resistant human breast cancer variant which retains sensitivity to the steroidal antiestrogen ICI 182, 780
Cancer Res.
MCF7/LCC9: an antiestrogen resistant MCF-7 variant in which acquired resistance to the steroidal antiestrogen ICI 182, 780 confers an early crossresistance to the non-steroidal antiestrogen tamoxifen
Cancer Res.
Antiestrogen resistance in breast cancer and the role of estrogen receptor signaling
Oncogene
Differential effects of estrogen and antiestrogen on in vitro clonogenic growth of human breast cancers in soft agar
J. Natl. Cancer Inst.
Influence of synthetic oestrogens upon advanced malignant disease
Br. Med. J.
Synthetic oestrogens in mammary cancer
Lancet
Tamoxifen and 17β-estradiol reduce the membrane fluidity of human breast cancer cells
J. Natl. Cancer Inst.
Bcl-2 inhibition of autophagy: a new route to cancer?
Cancer Res.
Apoptosis and genomic instability
Nat. Rev. Mol. Cell Biol.
The role of apoptosis in cancer development and treatment response
Nat. Rev. Cancer
The coordinate release of cytochrome c during apoptosis is rapid, complete and kinetically invariant
Nat. Cell Biol.
Role of Bcl-2 family proteins in a non-apoptotic programmed cell death dependent on autophagy genes
Nat. Cell Biol.
BID-deficient breast cancer MCF-7 cells as a model for the study of autophagy in cancer therapy
Autophagy
The evolutionarily conserved domain of Beclin 1 is required for Vps34 binding, autophagy and tumor suppressor function
Autophagy
Induction of autophagy and inhibition of tumorigenesis by beclin 1
Nature
Cell death independent of caspases: a review
Clin. Cancer Res.
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Lecture presented at the ‘18th International Symposium of the Journal of Steroid Biochemistry and Molecular Biology’, 18–21 September 2008, Seefeld, Tyrol, Austria.