Adaptive Hypersensitivity Research Articles

Mechanistic Effects of Estrogens on Breast Cancer.

Current concepts regarding estrogen and its mechanistic effects on breast cancer in women are evolving. This article reviews studies that address estrogen-mediated breast cancer development, the prevalence of occult tumors at autopsy, and the natural history of breast cancer as predicted by a newly developed tumor kinetic model. This article reviews previously published studies from the authors and articles pertinent to the data presented. We discuss the concepts of adaptive hypersensitivity that develops in response to long-term deprivation of estrogen and results in both increased cell proliferation and apoptosis. The effects of menopausal hormonal therapy on breast cancer in postmenopausal women are interpreted based on the tumor kinetic model. Studies of the administration of a tissue selective estrogen complex in vitro, in vivo, and in patients are described. We review the various clinical studies of breast cancer prevention with selective estrogen receptor modulators and aromatase inhibitors. Finally, the effects of the underlying risk of breast cancer on the effects of menopausal hormone therapy are outlined. The overall intent of this review is to present data supporting recent concepts, discuss pertinent literature, and critically examine areas of controversy.

Management of Aromatase Inhibitor-Resistant Disease with Estrogen, Selective Estrogen Receptor Down-Regulators, and Other Agents

With increased use of aromatase inhibitors (AIs) in the adjuvant and first-line metastatic settings for postmenopausal women with estrogen receptor (ER)-positive breast cancer, systemic relapse or progressive metastatic disease on AIs is an increasingly encountered clinical scenario. Overcoming resistance is a priority. Patients with AI-resistant disease represent a heterogeneous population, with diverse mechanisms of resistance dictating varied sensitivity to subsequent treatment. Cells with persistent dependence on ER signaling may be inhibited by fulvestrant, in which case dose-dependent ER downregulation and activity favor high-dose, loading schedule fulvestrant. In direct contrast, cells with long-term estrogen deprivation with adaptive estrogen hypersensitivity may be inhibited by exposure to estrogen. Cell survival by alternate growth signaling pathways may be inhibited by targeted agents. Currently missing, however, are predictive biomarkers to identify underlying resistance mechanisms and guide effective post-AI therapy for individual patients.

Biologic Basis of Sequential and Combination Therapies for Hormone-Responsive Breast Cancer

Although pharmacologic therapies that reduce or block estrogen signaling are effective treatments of estrogen receptor (ER)-positive breast cancer, acquired resistance to individual drugs can develop. Furthermore, this approach is ineffective as initial therapy for a subgroup of receptor-positive patients. The mechanisms of drug resistance are not completely understood, but the presence of alternative signaling pathways for activating ER response appears to play a significant role. Cross-talk between signaling pathways can activate ERs when conventional ER pathways are blocked or inactivated. For example, signaling via epidermal growth factor or HER-2 receptors, mitogen-activated protein kinases, phosphatidylinositol 3' kinase/protein kinase B, and vascular endothelial growth factor receptor can lead to estrogen-independent stimulation of ERs and tumor growth. The discovery that alternative pathways are involved in estrogen signaling has prompted development of newer endocrine therapies, such as aromatase inhibitors and pure estrogen antagonists, with distinct mechanisms for interrupting signal transduction. The existence of multiple pathways may explain the effectiveness of follow-up therapy with a different class of endocrine agents after failure of prior endocrine treatment. Because they do not have the partial agonist activity of tamoxifen that is enhanced by the adaptive hypersensitivity process, these alternative endocrine agents may play an increasingly important role in the treatment of ER-positive breast cancer. Although optimal sequencing of these agents has not been determined and is continuing to evolve, current evidence allows rational recommendations to be made. The multiple pathways involved in activating ERs also provide a rationale for combining endocrine and non-endocrine therapies that block different signaling pathways, which may have synergistic and overlapping interactions.

The Phenomenon of the Switching of Estrogen Effects and Joker Function of Glucose: Similarities and Relation to Age‐Associated Pathology and Approaches to Correction

Estrogens and glucose are characterized by a myriad of functions that can be reduced to a small number of principal actions. In aging there is a simultaneous increase in the prevalence of diseases connected with estrogen deficiency as well as with estrogenic excess and associated with the phenomenon of the switching of estrogen effects (PSEE). Estrogens possess hormonal and genotoxic properties. An increase in genotoxic effect (isolated or combined with a decrease in hormonal effect) can influence the course of age-associated diseases that, contrary to the situation with adaptive hypersensitivity to estrogens, may become less favorable or more aggressive. Inductors of PSEE include smoking, irradiation, and aging. Yet with "glycemic load" and the endocrine effect of glucose (the stimulation of insulin secretion), reactive oxygen species are formed in multiple sites, including adipose tissue. The ratio between hormonal and genotoxic effects reflects a "joker" function of glucose and can be conditioned by endogenous (perhaps including genetic) and exogenous factors. The shift in this glucose-associated ratio may selectively encourage some chronic non-communicable diseases. Several groups of treatments can be distinguished including alleviators of PSEE and insulin resistance syndrome (biguanides, glitazones, statins, modifiers of adipocytokines secretion, etc.) as well as other compounds aimed to optimally orchestrate the balance between endocrine and DNA-damaging effects of estrogens and glucose.

Long-term estradiol deprivation in breast cancer cells up-regulates growth factor signaling and enhances estrogen sensitivity

Deprivation of estrogen causes breast tumors in women to adapt and develop enhanced sensitivity to this steroid. Accordingly, women relapsing after treatment with oophorectomy, which substantially lowers estradiol for a prolonged period, respond secondarily to aromatase inhibitors with tumor regression. We have utilized in vitro and in vivo model systems to examine the biologic processes whereby long-term estradiol deprivation (LTED) causes cells to adapt and develop hypersensitivity to estradiol. Several mechanisms are associated with this response, including up-regulation of estrogen receptor-alpha (ERalpha) and the MAP kinase, phosphoinositol 3 kinase (PI3-K) and mammalian target of rapamycin (mTOR) growth factor pathways. ERalpha is four- to tenfold up-regulated and co-opts a classical growth factor pathway using Shc, Grb-2 and Sos. This induces rapid non-genomic effects which are enhanced in LTED cells. The molecules involved in the non-genomic signaling process have been identified. Estradiol binds to cell membrane-associated ERalpha, which physically associates with the adaptor protein Shc, and induces its phosphorylation. In turn, Shc binds Grb-2 and Sos, which result in the rapid activation of MAP kinase. These non-genomic effects of estradiol produce biologic effects as evidenced by Elk-1 activation and by morphologic changes in cell membranes. Additional effects include activation of the PI3-K and mTOR pathways through estradiol-induced binding of ERalpha to the IGF-I and epidermal growth factor receptors. A major question is how ERalpha locates in the plasma membrane since it does not contain an inherent membrane localization signal. We have provided evidence that the IGF-I receptor serves as an anchor for ERalpha in the plasma membrane. Estradiol causes phosphorylation of the adaptor protein, Shc and the IGF-I receptor itself. Shc, after binding to ERalpha, serves as the 'bus' which carries ERalpha to Shc-binding sites on the activated IGF-I receptors. Use of small inhibitor (si) RNA methodology to knockdown Shc allows the conclusion that Shc is needed for ERalpha to localize in the plasma membrane. In order to abrogate growth factor-induced hypersensitivity, we have utilized a drug, farnesylthiosalicylic acid, which blocks the binding of GTP-Ras to its membrane acceptor protein, galectin 1, and reduces the activation of MAP kinase. We have also shown that this drug is a potent inhibitor of mTOR as an additional mechanism of inhibition of cell proliferation. The concept of 'adaptive hypersensitivity' and the mechanisms responsible for this phenomenon have important clinical implications. The efficacy of aromatase inhibitors in patients relapsing on tamoxifen could be explained by this mechanism and inhibitors of growth factor pathways should reverse the hypersensitivity phenomenon and result in prolongation of the efficacy of hormonal therapy for breast cancer.

Adaptive hypersensitivity to estrogen: Mechanisms and clinical relevance to aromatase inhibitor therapy in breast cancer treatment

Breast tumors in women can adapt to endocrine deprivation therapy by developing hypersensitivity to estradiol. For this reason, aromatase inhibitors can be effective in women relapsing after treatment with tamoxifen or following oophorectomy. To understand the mechanisms responsible, we examined estrogenic stimulation of cell proliferation in a model system and provided in vitro and in vivo evidence that long-term estradiol deprivation (LTED) causes “adaptive hypersensitivity”. The primary mechanisms responsible involve up-regulation of ERα as well as the MAP kinase, PI-3 kinase, and mTOR growth factor pathways. ERα is 4–10-fold up-regulated and co-opts a classical growth factor pathway using Shc, Grb2, and Sos. This induces rapid non-genomic effects which are enhanced in LTED cells. Estradiol binds to cell membrane associated ERα, physically associates with the adaptor protein Shc, and induces its phosphorylation. In turn, Shc binds Grb2 and Sos which result in the rapid activation of MAP kinase. These non-genomic effects of estradiol produce biologic effects as evidenced by Elk activation and by morphologic changes in cell membranes. Additional effects include activation of PI-3 kinase and mTOR pathways through estradiol induced binding of ERα to the IGF-1 and EGF receptors. Further proof of the non-genomic effects of estradiol involved use of “designer” cells which selectively express ERα in nucleus, cytosol, and cell membrane. We have used a new downstream inhibitor of these pathways, farnesyl-thio-salicylic acid (FTS), to block proliferation in hypersensitive cells as a model for a potentially effective strategy for treatment of patients.

Proceedings of the Fourth International Conference on Recent Advances and Future Directions in Endocrine Manipulation of Breast Cancer: Conference Summary Statement

Proceedings of the Fourth International Conference on Recent Advances and Future Directions in Endocrine Manipulation of Breast Cancer: Conference Summary Statement

Adaptive hypersensitivity to estrogen: mechanism for sequential responses to hormonal therapy in breast cancer.

Clinical observations demonstrate that women with breast cancer often respond to subsequent endocrine manipulation after resistance to initial hormonal therapy develops. As a mechanistic explanation for these findings, we hypothesized that human breast tumors can adapt in response to the pressure exerted by endocrine therapy with development of hypersensitivity to estradiol. To understand the signaling pathways responsible, we examined estrogenic stimulation of cell proliferation in a model system and provided in vitro and in vivo evidence that long-term deprivation of estradiol (LTED) causes adaptive hypersensitivity. Even though the estrogen receptor alpha (ERalpha) is markedly up-regulated in LTED cells, the enhanced responses to estradiol do not appear to involve mechanisms acting at the level of transcription of estrogen-regulated genes. We found that ERalpha co-opts a classical growth factor pathway and induces rapid nongenomic effects that are enhanced in LTED cells. Estradiol binds to cell membrane-associated ERs, physically associates with the adaptor protein Shc, and induces its phosphorylation. In turn, Shc binds Grb2 and Sos, which result in the rapid activation of mitogen-activated protein kinase. These nongenomic effects of estradiol produced biological effects, as evidenced by Elk-1 activation and by morphological changes in cell membranes. The mechanistic pathways involved in adaptive hypersensitivity suggest that inhibitors of the mitogen-activated protein kinase and phosphatidylinositol-3-OH kinase pathways might prevent the development of adaptive hypersensitivity and allow more prolonged efficacy of endocrine therapies.

Adaptive hypersensitivity following long-term estrogen deprivation: involvement of multiple signal pathways.

Long-term estrogen deprivation causes hypersensitivity of MCF-7 cells to the mitogenic effect of estradiol (E2) which is associated with activation of mitogen-activated protein kinase (MAPK). However, several lines of evidence indicate that MAPK activation is not the exclusive mechanism for E2 hypersensitivity and multiple signal pathways might be involved. The current study explores the possible role of the PI3 kinase (PI3K) pathway in development of E2 hypersensitivity. Basal PI3K activity in long-term estrogen deprived MCF-7 cells (LTED) was elevated as evidenced by increased phosphorylation of three downstream effectors, Akt, p70 S6 kinase, and eukaryotic initiation factor-4E binding protein (4E-BP1), which was blocked by the specific inhibitor of PI3K, LY294002. Dual blockade of both MAPK and PI3K completely reversed E2 hypersensitivity of LTED cells. Enhancement in aromatase activity is another phenomenon accompanied with E2 hypersensitivity. In aromatase over-expressing MCF-7 cells, aromatase activity was reduced by inhibitors of MAPK and PI3K suggesting the involvement of protein phosphorylation in the regulation of aromatase activity. Our data suggest that in addition to the MAP kinase pathway, activation of the PI3 kinase pathway is involved in E2 hypersensitivity, which develops during adaptation of MCF-7 cells to the low estrogen environment.

Adaptive hypersensitivity to estrogen: mechanism for superiority of aromatase inhibitors over selective estrogen receptor modulators for breast cancer treatment and prevention.

Clinical observations suggest that human breast tumors can adapt to endocrine therapy by developing hypersensitivity to estradiol (E(2)). To understand the mechanisms responsible, we examined estrogenic stimulation of cell proliferation in a model system and provided in vitro and in vivo evidence that long-term E(2) deprivation (LTED) causes "adaptive hypersensitivity". The enhanced responses to E(2) do not involve mechanisms acting at the level of transcription of estrogen-regulated genes. We found no evidence of hypersensitivity when examining the effects of E(2) on regulation of c-myc, pS2, progesterone receptor, several estrogen receptor (ER) reporter genes, or c-myb in hypersensitive cells. Estrogen deprivation of breast cells long-term does up-regulate both the MAP kinase and phosphatidyl-inositol 3-kinase pathways. As a potential explanation for up-regulation of these signaling pathways, we found that ERalpha is 4- to 10-fold up-regulated and co-opts a classic growth factor pathway using Shc, Grb-2 and Sos. This induces rapid non-genomic effects which are enhanced in LTED cells. E(2) binds to cell membrane-associated ERalpha, physically associates with the adapter protein SHC, and induces its phosphorylation. In turn, Shc binds Grb-2 and Sos, which results in the rapid activation of MAP kinase. These non-genomic effects of E(2) produce biological effects as evidenced by Elk activation and by morphological changes in cell membranes. Further proof of the non-genomic effects of E(2) involved use of cells which selectively expressed ERalpha in the nucleus, cytosol and cell membrane. We created these COS-1 "designer cells" by transfecting ERalpha lacking a nuclear localization signal and containing a membrane localizing signal. The concept of "adaptive hypersensitivity" and the mechanisms responsible for this phenomenon have important clinical implications. Adaptive hypersensitivity would explain the superiority of aromatase inhibitors over the selective ER modulators (SERMs) for treatment of breast cancer. The development of highly potent third-generation aromatase inhibitors allows reduction of breast tissue E2 to very low levels and circumvents the enhanced sensitivity of these cells to the proliferative effects of E(2). Clinical trials in the adjuvant, neoadjuvant and advanced disease settings demonstrate the greater clinical efficacy of the aromatase inhibitors over the SERMs. More recent observations indicate that the aromatase inhibitors are superior for the prevention of breast cancer as well. These observations may be explained by the hypothesis that estrogens induce breast cancer both by stimulating cell proliferation and by their metabolism to genotoxic products. The SERMs block ER-mediated proliferation only, whereas the aromatase inhibitors exert dual effects on proliferation and genotoxic metabolite formation.

Adaptive mechanisms induced by long-term estrogen deprivation in breast cancer cells

Clinical observations suggest that human breast tumors can adapt in response to endocrine therapy by developing hypersensitivity to estradiol. To understand the mechanisms responsible, we examined estrogenic stimulation of cell proliferation in a model system and provided evidence that long-term deprivation of estradiol causes adaptive hypersensitivity. The enhanced responses to estradiol do not involve mechanisms acting at the level of transcription of estrogen regulated genes. We found no evidence of hypersensitivity when examining the effects of estradiol on regulation of c-myc, pS2, progesterone receptor, several ER reporter genes or c-myb in hypersensitive cells. On the other hand, deprivation of breast cells long term was found to up-regulate a separate pathway whereby the estrogen receptor co-opts a classical growth factor pathway and induces rapid non-genomic effects. Through this pathway, estradiol caused rapid activation of mitogen-activated protein (MAP) kinase. In exploring the mechanisms mediating this event, we found that estradiol binds to cell membrane associated estrogen receptors and causes phosphorylation of Shc, an adaptor protein usually involved in growth factor signaling pathways. ERα was found to complex with Shc under these conditions. In turn, Shc bound Grb-2 and Sos which resulted in the activation of MAP kinase. The pure antiestrogen, ICI 182,780, blocked several steps in the rapidly responding ER alpha, Shc, MAP kinase pathway. These non-genomic effects of estradiol produced biologic effects by activating Elk and by inducing morphologic changes in cell membranes. Using confocal microscopy, we demonstrated that estradiol caused a rapid alteration in membrane ruffling, the formation of pseudopodia and translocation of ER alpha to regions contiguous with the cell membrane. These morphologic effects could be blocked with a pure anti-estrogen. We conclude that long-term estradiol deprived cells utilize both genomic (transcriptional) and rapid, non-genomic estradiol induced pathways. We postulate that synergy between these two pathways acting at the level of the cell cycle is responsible for adaptive hypersensitivity.

Adaptive hypersensitivity to estradiol: potential mechanism for secondary hormonal responses in breast cancer patients

Women with hormone dependent breast cancer initially respond to hormone deprivation therapy with tamoxifen or oophorectomy for 12–18 months but later relapse. Upon secondary therapy with aromatase inhibitors, patients often experience further tumor regression. The mechanisms responsible for secondary responses are unknown. We postulated that hormone deprivation induces hypersensitivity to estradiol. Evidence of this phenomenon was provided in a model system involving MCF-7 cells grown in vitro and in xenografts. To determine if the ER transcriptional process is involved in hypersensitivity, we examined the effect of estradiol on ER reporter activity, PgR, PS2, and c-myc as markers and found no alterations in hypersensitive cells. Next, we examined whether MAP kinase may be upregulated in the hypersensitive cells as a reflection of increased growth factor secretion or action. Basal MAP kinase activity was increased both in vitro and in vivo in hypersensitive cells. Proof of principle studies indicated that an increase in MAP kinase activity induced by TGFα administration caused a two- to three-fold shift to the left in estradiol dose response curves in wild type cells. Blockade of MAP kinase with PD98059 returned the shifted curve back to baseline. These data suggested that MAP kinase overexpression could induce hypersensitivity. To determine why MAP kinase was increased, we excluded constitutive receptor activity and growth factor secretion by the demonstration that the pure anti-estrogen, ICI 182780, could inhibit MAP kinase activation. We also excluded hypersensitivity to estradiol induced growth factor secretion, and thus MAP kinase activation, since estradiol stimulated MAP kinase at 24, 48, and 72 h at the same concentrations in hypersensitive as in wild type cells. Surprisingly, a series of experiments suggested that MAP kinase increased in hypersensitive cells as a result of estrogen activation via a non-genomic pathway. We examined the classical signal pathway in which SHC is phosphorylated and binds to SOS and GRB-2 to activate Ras, Raf, and MAP kinase. With 5–20 min of exposure, estradiol caused binding of SHC to the estrogen receptor, phosphorylation of SHC, binding of GRB-2 to SOS, and activation of MAP kinase. All of these affects could be blocked by ICI 182780. Taken together, these observations suggest that the cell membrane ER pathway may be responsible for upregulation of MAP kinase and hypersensitivity in cells adapted to estradiol deprivation.

Use of aromatase inhibitors in breast carcinoma.

Aromatase, a cytochrome P-450 enzyme that catalyzes the conversion of androgens to estrogens, is the major mechanism of estrogen synthesis in the post-menopausal woman. We review some of the recent scientific advances which shed light on the biologic significance, physiology, expression and regulation of aromatase in breast tissue. Inhibition of aromatase, the terminal step in estrogen biosynthesis, provides a way of treating hormone-dependent breast cancer in older patients. Aminoglutethimide was the first widely used aromatase inhibitor but had several clinical drawbacks. Newer agents are considerably more selective, more potent, less toxic and easier to use in the clinical setting. This article reviews the clinical data supporting the use of the potent, oral competitive aromatase inhibitors anastrozole, letrozole and vorozole and the irreversible inhibitors 4-OH androstenedione and exemestane. The more potent compounds inhibit both peripheral and intra-tumoral aromatase. We discuss the evidence supporting the notion that aromatase inhibitors lack cross-resistance with antiestrogens and suggest that the newer, more potent compounds may have a particular application in breast cancer treatment in a setting of adaptive hypersensitivity to estrogens. Currently available aromatase inhibitors are safe and effective in the management of hormone-dependent breast cancer in post-menopausal women failing antiestrogen therapy and should now be used before progestational agents. There is abundant evidence to support testing these compounds as first-line hormonal therapy for metastatic breast cancer as well as part of adjuvant regimens in older patients and quite possibly in chemoprevention trials of breast cancer.