Intracrinology of estrogens and androgens in breast carcinoma

Abstract

Intratumoral metabolism and synthesis of biologically active steroids such as estradiol and 5α-dihydrotestosterone as a result of interactions of various enzymes are considered to play very important roles in the pathogenesis and development of hormone-dependent breast carcinoma. Among these enzymes involved in estrogen metabolism, intratumoral aromatase play an important role in converting androgens to estrogens in situ from serum and serving as the source of estrogens, especially in postmenopausal patients with breast carcinoma. However, other enzymes such as 17β-hydroxysteroid dehydrogenase (17β-HSD) isozymes, estrogen sulfatase (STS), and estrogen sulfotransferase, which contribute to in situ availability of biologically active estrogens, also play pivotal roles in this intratumoral estrogen production above. Androgen action on human breast carcinoma has not been well-studied but are considered important not only in hormonal regulation but also other biological features of carcinoma cells. Intracrine mechanisms also play important roles in androgen actions on human breast carcinoma cells. Among the enzymes involved in biologically active androgen metabolism and/or synthesis, both 17β-hydroxysteroid dehydrogenase type 5 (17βHSD5; conversion from circulating androstenedione to testosterone) and 5α-reductase (5αRed; reduction of testosterone to DHT (5α-dihydrotestosterone) were expressed in breast carcinoma tissues, and in situ production of DHT has been proposed in human breast cancer tissues. However, intracrine mechanisms of androgens as well as their biological or clinical significance in the patients with breast cancer have not been fully elucidated in contrast to those in estrogens.

Introduction

Biologically active hormones, whether peptides or steroids, are synthesized and secreted from the endocrine organs such as adrenal cortex, or ovary, or pituitary glands. These hormones are transported through the circulation, and act on their target tissues where their specific receptors are expressed (Fig. 1). This system of hormone actions has been called “endocrine”, and various biological/clinical features of endocrine target tissues are well-known to be influenced by plasma concentration of the biologically active hormones. Therefore, in the fields of endocrinology, it is very important to evaluate serum or urinary concentrations of hormones in order to obtain a better understanding of physiology and pathology of hormones actions. These locally produced hormones can also act in the same cell (autocrine) or neighboring cells (paracrine) without their release into the circulation.

However, it is also true that a large proportion of androgens in men (approximately 50%) and estrogens in women (approximately 75% before menopause, and close to 100% after menopause) were produced in peripheral hormone-target tissues from abundantly present circulating precursor steroids [1], where the enzymes involved in the formation of androgens and estrogens are expressed (Fig. 1). These locally produced bioactive androgens and/or estrogens exert their action in the cells where synthesis occurs without release in the extracellular space including circulation. This phenomenon is different from the classical concept of endocrinology such as autocrine, paracrine, and endocrine. This mechanism has been termed “intracrine”. It is Labrie and colleagues who elegantly described the formation of active androgens such as dehydrotestosterone (DHT) from the inactive adrenal precursors, dehydroepiandrosterone (DHEA), (DHEA-S (sulfate)) and/or androstenedione locally in the some tissues or cells in adenocarcinoma of the prostate where biosynthesis takes place without release into the extracellular space as “intracrine activity” [1], [2].

It then becomes very important to evaluate physiological and/or pathological significance of this intracrine activity compared to endocrine activity. In classical endocrine systems, among those produced and secreted from the endocrine organs, only a small amount of hormones secreted is in general utilized in the target tissues or exerts their effects. The great majority of these hormones is actually metabolized or converted to inactive forms. In contrast, an intracrine system requires minimal amounts of biologically active hormones to exert their maximum hormonal effects. Therefore, the intracrine system is considered a markedly efficient mode of hormone action and plays an important role, especially in the development of hormone-dependent neoplasms including human prostate, breast, endometrial, and ovarian malignancies. It is also important to note that, in an intracrine system, serum concentrations of hormones do not necessarily reflect the local hormonal activities in the target tissues. Therefore, it becomes very important to study how the hormones are metabolized and/or synthesized in the tissue where they exert their actions.

In this review, we summarize intratumoral production of sex steroids including estrogens and androgens in human breast carcinoma tissues, and discuss the potential biological and/or clinical significance of intratumoral production of sex steroids in these carcinomas.

The great majority of human breast carcinomas express estrogen receptor (ER) in carcinoma or parenchymal cells. These cases are termed hormone- or estrogen-dependent breast carcinoma, and estrogens, especially 17β-estradiol (E2), a biologically potent estrogen, contribute greatly to the growth and development of carcinoma cells and some of these carcinoma cases actually require estrogens for their continued growth and other biological behaviors [3].

It then becomes very important to determine the possible sources of these estrogens that influence various biological behaviors of breast cancers. It is well-known that estradiol originated from different sources before and after the menopause in women. In premenopausal women, the ovary or membrana granulosa of dominant follicles is the main source of abundant circulating estrogens [4], [5]. However, as mentioned above after menopause, estrogens are produced primarily through conversion of androgens of both adrenal and ovarian origins, especially of zona reticularis origin of adrenal cortex [6]. The conversion of androgens to estrone occurs principally in peripheral tissues, including skin [7], muscle [8], fat [8], and bone [9]. This conversion is catalyzed by the aromatase enzyme complex [3], [5]. However, the great majority of estrone in circulation, including postmenopausal women, is present as sulfated form or estrone sulfate (E1-S) and steroid sulfatase (STS) hydrolyzes circulating E1-S to E1 in various human tissues [10], [11]. Estrogen sulfotransferase (EST) (SULT 1E1 or STE gene) is a member of the superfamilly of steroid-sulfotransferases, and sulfonates estrogens to biologically inactive estrogen sulfates [12], [13], [14]. Therefore, EST and STS play very important roles in maintaining an availability of biologically active estrogens in the tissues. Estrone is subsequently reduced to 17β-estradiol by 17β-hydroxysteroid dehydrogenase (HSD) type 1, which is also widely distributed in various peripheral tissues [15], [16], [17].

Increased peripheral conversion of androgens to estrogens may result in elevated serum levels of estrogens. Therefore, numerous studies have been performed to examine the subtle differences of serum estrogen concentrations between breast cancer patients and their age matched control population. Several epidemiological studies did indicate that plasma estradiol, adrenal androgens, and testosterone levels are higher in women who will develop neoplasms over a period of several years than in those who do not [18]. However, results of other studies [19], [20] were not necessarily consistent with those above.

Miller et al. [21] and Perel et al. [22] both independently demonstrated that human breast and its neoplasms can produce 17β-estradiol in vitro. Yue et al. subsequently reported that in situ synthesis of estrogen predominates over uptake from plasma as a means of maintaining breast tissue estradiol concentrations after menopause [23]. These findings clearly indicate the biological importance of elevated in situ estrogen concentrations as a result of intratumoral estrogen production in postmenopausal human breast cancer patients. The pathways or cascades of intratumoral estrogen production in human breast cancer tissue is summarized in Fig. 2. It is true that the biological significance of in situ estrogen production still remains controversial with regard to development and biological behavior of breast cancer but an increasing number of studies have indicated that in patients with estrogen-dependent breast carcinoma, especially in postmenopausal women, intratumoral estrogens derived from in situ aromatization could function as an autocrine growth and a mitogenic factor and could impart a growth advantage to these cancer cells, regardless of serum concentration of estrogens. Therefore, estrogen-dependent breast carcinoma in which in situ conversions from serum androgen to biologically active estrogens occur should also be considered as “intracrine” tissues [24].

AR or androgen receptor is expressed in a majority of human breast carcinoma tissues, especially in carcinoma cells (Fig. 3), more widely than estrogen receptor [25], [26], [27], [28]. In addition, Farmer et al. recently identified the group of breast carcinoma cases with increased androgen signaling and some apocrine features in a microarray study [29]. These tumors are ER negative and retention of androgenic signals may further subclassify ER negative breast carcinoma into subtypes which may respond to various target specific therapy. These findings all suggest potential important roles of androgens in human breast carcinomas. However, the possible effects or actions of androgens on human breast carcinoma cells have not necessarily been well-examined.

The effects of androgens are considered to predominantly exert anti-proliferative effects via androgen receptor or AR in human breast carcinoma cells [25], [26], [27], [28], although some divergent or conflicting findings have been reported. AR is expressed in a majority of human breast carcinoma tissues [30], [31], [32], [33], suggesting important roles of androgens in breast carcinomas. 5α-Dihydrotestosterone (DHT) binds with the highest affinity to AR, and together with testosterone promotes AR transcriptional activity [34]. Androgen concentrations have been previously examined in breast cancers by two groups [35], [36], and the potent androgen DHT was demonstrated to be significantly higher in breast carcinoma tissues than in plasma in these studies. Androgen-producing enzymes, such as 17β-hydroxysteroid dehydrogenase type 5 (17βHSD5; conversion from circulating androstenedione to testosterone) and 5α-reductase (5αRed; reduction of testosterone to DHT) were expressed in breast carcinoma tissues [33], and in situ production of DHT has been proposed in breast cancer tissues [34]. The enzymes involved in in situ androgen production in human breast carcinoma is summarized in Fig. 4.

Androgen actions are mediated through an interaction with androgens and AR. Therefore, it becomes very important to evaluate both AR expression and intratumoral DHT concentration in the breast carcinoma tissues, in order to obtain a better understanding of the androgenic actions.

Correlation between AR status and clinical outcome of breast carcinoma patients has been examined by several groups, but the results were not necessarily consistent. Bryan et al. [35] found a significant association between AR status evaluated by AR assays and overall survival of the patients. Soreide et al. [30], however, did not detect any significant correlation between AR status and relapse-free survival. In more recent study, Kuenen-Boumeester et al. [32] performed immunohistochemistry for AR in 153 breast carcinomas, and reported that AR status was a significant prognostic factor for disease-free survival, but was not an independent factor following a multiple analysis. If DHT is involved in growth inhibition through AR, residual cancer cells following surgical treatment in AR and 5αRed1 double-positive breast carcinomas possibly grow slowly in the presence of locally produced DHT, which may subsequently result in a better clinical outcome of these patients. Inconsistent results regarding the correlation between AR status and prognosis in previous studies may partly be due to different ratios of breast carcinomas positive for both AR and 5αRed1 examined. Previous studies demonstrated a significant association of intratumoral concentrations between DHT and testosterone in breast carcinomas, suggesting that the intratumoral DHT concentrations were influenced by amounts of the precursor [35], [36]. Aromatase catalyzes the conversion of androstenedione and testosterone, which are precursors of DHT, to estrone and estradiol, respectively. However, DHT itself is nonaromatizable. Previously, Spinola et al. reported that treatment with an aromatase inhibitor (4-hydroxyandrostenedione) markedly elevated intratumoral testosterone concentrations in dimethylbenz(a)anthracene (DMBA)-induced rat mammary tumors [36], and Sonne-Hansen and Lykkesfeldt [37] recently showed that aromatase preferred testosterone as a substrate in MCF-7 cells. In addition, the aromatase inhibitor letrozole was demonstrated to block conversion of androgens into estrogens with a subsequent increment of intraovarian androgens [38], [39]. Therefore, aromatase is suggested a negative regulator for in situ production of DHT in breast carcinoma tissues by possibly reducing concentrations or availability of the precursor testosterone. Administration of androgens combined with anti-estrogen has been more effective than that of anti-estrogen alone in breast cancer patients, and the additive inhibitory effects were exerted in part by different mechanisms [40]. In addition, results of recent treatments, such as anastrolozole letrozole and exemestane, compared to anti-estrogen tamoxifen [40], [41], [42], although it might be due to agonistic effects of tamoxifen in estrogen-deprived environment [43]. It awaits further investigations to clarify the correlation between in situ androgen production or intracrinology of androgens and androgen actions in human breast carcinoma cells.