An overview of the world literature on spontaneous tumors in nonhuman primates

A review of the world literature has revealed a total of 783 spontaneous neoplasms that have occurred in nonhuman primates, all similar to tumors in human beings. Carcinomas were the most frequently reported. Neoplasms occurred most frequently in older animals as they generally do in man. There were more reports on neoplasms in baboons and macaques than in other species. The digestive system was more commonly affected than were other organ systems.     

Expression of sex steroid receptors and IGF-1 mRNA in breast tissue--effects of hormonal treatment

The mechanisms behind increased breast tissue proliferation and a possibly increased breast cancer risk in women using hormonal contraception (HC) and hormonal replacement therapy (HRT) are incompletely understood. We analyzed breast tissue from 20 premenopausal and seven postmenopausal women undergoing reduction mammoplasties for estrogen receptor (ER) and progesterone receptor (PR) content as well as mRNA levels for ER, PR and insulin-like growth factor-1 (IGF-1). The receptor values were correlated to IGF-1 mRNA concentrations and levels of steroid and peptide hormones and SHBG. In women using HC, we found significantly lower ER values (p=0.02) but non-significantly lower ER mRNA levels compared to those in naturally cycling women. PR and PR mRNA were no different. Women on HC displayed a higher breast tissue proliferation (p=0.05) expressed as Ki-67, MIB-1 positivity, which was correlated with IGF-1 mRNA (r_s=0.82, p=0.04). Since the concentration of sex steroid receptors in breast tissue is comparatively low and steroid receptors are down-regulated during hormonal treatment, mechanisms other than direct sex steroid receptor action are likely to be present. Our results suggest a role for IGF-1 in the proliferative response of breast tissue during exogenous hormonal treatment.     

Topographic recognition of cyclic hydrocarbons and related compounds by receptors for androgens, estrogens, and glucocorticoids

The structural requirements for the interaction of about 80 cyclic hydrocarbons and related compounds with the androgen receptor of rat ventral prostate, the estrogen receptor of human breast tumor MCF-7 cells, and the glucocorticoid receptor of rat liver were examined by comparing their abilities to compete with radioactive hormones for binding to the respective receptors. The results indicate that the receptor-binding affinity of a compound is dependent on its electronic configuration and geometrical similarity to a portion of a natural steroid hormone which can be recognized by local ligand-binding sites in the receptor. For the estrogen receptor, beta-phenols are more active than the corresponding alpha-phenols, whereas nonphenolic compounds are totally inactive. For androgen and glucocorticoid receptors, alpha-phenols are more active than beta-phenols. The androgen receptor can interact stereospecifically with nonoxygenated and nonalkylated cyclic hydrocarbons, such as 10,11-dihydro-5H-dibenzo[a,d] cycloheptene or 9,10-dihydrophenanthrene, which can, in vivo, inhibit the androgen-dependent growth of the male accessory reproductive organs. The affinities of naphthalene, anthracene, phenanthrene, biphenyl, and adamantane toward glucocorticoid and androgen receptors can be enhanced by acetylation or ethanolization of these ligands. Our results also indicate that, while the hormonal action of a steroid may be dependent on the interaction of a functional group on the hormone with a specific group on the receptor, the presence of such a group may not be required for the antagonistic activity of a compound that can physically block hormone binding to the receptor. Thus, many small molecules that were hitherto considered to be biologically inert may interact with steroid receptors specifically and affect hormonal activities in vivo.     

A multidrug-resistant MCF-7 human breast cancer cell line which exhibits cross-resistance to antiestrogens and hormone-independent tumor growth in vivo

MCF-7 human breast cancer cells provide a useful in vitro model system to study hormone-responsive breast cancer as they contain receptors for estrogen and progesterone, and estrogen both induces the synthesis of specific proteins in these cells and increases their rate of proliferation. An MCF-7 cell line which was selected for resistance to adriamycin (MCF-7/AdrR) exhibits the phenotype of multidrug resistance (MDR), and displays multiple biochemical changes. MDR in MCF-7/AdrR is also associated with a loss of mitogenic response to estrogen and the development of cross-resistance to the antiestrogen 4-hydroxytamoxifen. In addition, while the parental MCF-7 cell line responds to estrogen with increased levels of progesterone receptors and the secretion of specific proteins, these estrogen responses are lost in MCF-7/AdrR. Furthermore, while the formation of tumors in nude mice by wild-type MCF-7 cells is dependent upon the presence of estrogen, MCF-7/AdrR cells form tumors in the absence of exogenous estrogen administration. These changes in hormonal sensitivity and estrogen-independent tumorigenicity of the multidrug-resistant MCF-7 cell line are associated with a loss of the estrogen receptor and a concomitant increase in the level of receptors for epidermal growth factor. Thus, in MCF-7/AdrR cells, the development of MDR is associated with alterations in the expression of both cytosolic and membrane receptors, resulting in resistance to hormonal agents and the expression of hormone-independent tumor formation.     

Challenges to defining a role for progesterone in breast cancer

Progesterone is an ovarian steroid hormone that is essential for normal breast development during puberty and in preparation for lactation. The actions of progesterone are primarily mediated by its high affinity receptors, including the classical progesterone receptor (PR) -A and -B isoforms, located in diverse tissues such as the brain where progesterone controls reproductive behavior, and the breast and reproductive organs. Progestins are frequently prescribed as contraceptives or to alleviate menopausal symptoms, wherein progestin is combined with estrogen as a means to block estrogen-induced endometrial growth. Estrogen is undisputed as a potent breast mitogen, and inhibitors of the estrogen receptor (ER) and estrogen producing enzymes (aromatases) are effective first-line cancer therapies. However, PR action in breast cancer remains controversial. Herein, we review existing evidence from in vitro and in vivo models, and discuss the challenges to defining a role for progesterone in breast cancer.     

The DEK Oncogene Is a Target of Steroid Hormone Receptor Signaling in Breast Cancer

Expression of estrogen and progesterone hormone receptors indicates a favorable prognosis due to the successful use of hormonal therapies such as tamoxifen and aromatase inhibitors. Unfortunately, 15–20% of patients will experience breast cancer recurrence despite continued use of tamoxifen. Drug resistance to hormonal therapies is of great clinical concern so it is imperative to identify novel molecular factors that contribute to tumorigenesis in hormone receptor positive cancers and/or mediate drug sensitivity. The hope is that targeted therapies, in combination with hormonal therapies, will improve survival and prevent recurrence. We have previously shown that the DEK oncogene, which is a chromatin remodeling protein, supports breast cancer cell proliferation, invasion and the maintenance of the breast cancer stem cell population. In this report, we demonstrate that DEK expression is associated with positive hormone receptor status in primary breast cancers and is up-regulated in vitro following exposure to the hormones estrogen, progesterone, and androgen. Chromatin immunoprecipitation experiments identify DEK as a novel estrogen receptor α (ERα) target gene whose expression promotes estrogen-induced proliferation. Finally, we report for the first time that DEK depletion enhances tamoxifen-induced cell death in ER+ breast cancer cell lines. Together, our data suggest that DEK promotes the pathogenesis of ER+ breast cancer and that the targeted inhibition of DEK may enhance the efficacy of conventional hormone therapies.     

Estrogen and progesterone receptor proteins in breast cancer.

This article reviews the contribution of steroid hormone receptor studies to the resolution of a basic clinical problem: how to determine which cancers are hormone dependent without an actual treatment trial. Previously published studies on hormone receptor assays and analyses are reviewed, the current status of knowledge in the area is summarized in terms of its relevance to breast cancer cells, and future scenarios are proffered. Assay methods and available clinical results for cytoplasmic estrogen receptor identification in human breast cancer comprise the first section. Information on assaying nuclear estrogen receptor, and its relative clinical importance, is presented in Part 2. Assays to determine the presence of progesterone receptor in human breast cancer; the estogen regulation of such progesterone receptors; and clinical findings comprise Part 3. Studies of the mechanisms of estrogen action in the MCF-7 human breast cancer cell line are the subject of Part 4. Recent experiments in animals on the efficacy of antiestrogen treatments, such treatments in human in vitro cell cultures, and the few clinical trials available are presented in Part 5. It is emphasized that the simple presence or absence of estrogen receptors in tumors does not absolutely indicate whether growth of a particular tumor is sensitive to estrogens. Experimental appraoches, designed to further delineate this problem, are outlined, based on observations such as the finding that the probability of tumor regression correlates better with quantitative rather than with qualitative assessment of estrogen receptors; that the specific end product of hormone action is unknown and without this information an ideal biochemical marker to a tumor's sensitivity of hormones is unavailable; and that the complex sequence of biochemical events in the actions of estrogen needs more complete elucidation.     

An in vitro long-term culture model for normal human mammary gland: expression and regulation of steroid receptors

Steroids and their nuclear receptors play crucial roles in the development and maintenance of normal functions of the human mammary gland (HMG). They have also been implicated in breast carcinogenesis. However, the study of steroid action in normal HMG has been hampered by experimental difficulties. By using a newly established in vitro long-term culture method, we successfully cultured normal HMG tissue for more than 2 months without detriment to its morphology or steroid receptor expression. Expression of the cellular structural and extracellular matrix proteins was similar to that prior to culture, and HMG tissue retained its properties of steroid receptor expression and regulation. Addition of 17-beta estrogen to mammary tissues markedly increased the expression of progesterone receptor (PR) but only slightly affected that of the estrogen receptor (ER). Medroxyprogesterone acetate down-regulated the expression of PR within 24-48 h and also increased the expression of androgen receptor. When HMG tissue was cultured in medium containing normal or dextran-coated charcoal-stripped fetal calf serum or normal human serum, the expression and regulation of steroid hormone receptors were similar, although different in extent. When serum was omitted, the morphology of HMG was normal after 1 week, but the expression and regulation of ER and PR were altered. Thus, as HMGs retain the capacity to express steroid receptors in culture, this long-term culture system is probably a good model for studying the regulation of the mammary gland by steroids.     

Selective cytotoxicity of ascochlorin in ER-negative human breast cancer cell lines

While agents targeting estrogen receptors are most effective in adjuvant therapy for human breast cancers expressing estrogen receptors after surgery, breast cancers lacking estrogen receptor are clinically serious, because they are highly malignant and exhibit resistance to the usual anti-cancer drugs, including estrogen receptor-antagonists and DNA breaking agents. Here, we found that MX-1, a human breast cancer cell line lacking estrogen receptors, exhibited higher AP-1 activity and expressed higher levels of c-Jun, c-Fos, and Fra-1 when compared with conventional estrogen receptor-positive human breast cancer cell lines. The prenylphenol antibiotic ascochlorin suppressed the AP-1 activity of MX-1 cells, and selectively killed MX-1 cells, partly due to induction of apoptosis. Our results suggest that AP-1 is an effective clinical target molecule for the treatment of estrogen receptor-negative human breast cancer.     

Metabolism of pregnenolone by human breast cancer. Evidence for 17 alpha-hydroxylase and 17,20-lyase.

The metabolism of 7-(3)H-pregnenolone was studied in vitro using 16 human breast carcinomas. All mammary tumors transformed pregnenolone to progesterone. All estrogen receptor poor tumors and 4 out of 8 estrogen receptor rich tumors converted pregnenolone to 17-hydroxypregnenolone. Five estrogen receptor poor tumors showed the presence of 17,20-lyase as evidenced by formation of dehydroepiandrosterone and androstenedione. In two estrogen receptor poor tumors, conversions of pregnenolone to progesterone, 17-hydroxy pregnenolone, dehydroepiandrosterone, androstenedione and finally to estradiol was documented, providing a hypothetical pathway for steroid metabolism in human breast cancer. The conversion of pregnenolone to 17-hydroxypregnenolone was significantly less in receptor rich tumors and was totally absent in 4 receptor rich tumors with estrogen receptors of over 45 fmol/mg protein.     

Estradiol restores diabetes-induced reductions in sex steroid receptor expression and distribution in the vagina of db/db mouse model

Sex steroid hormones and receptors play an important role in maintaining vaginal physiology. Disruptions in steroid receptor signaling adversely impact vaginal function. Limited studies are available investigating the effects of diabetic complications on steroid receptor expression and distribution in the vagina. The goals of this study were to investigate type 2 diabetes-induced changes in expression, localization and distribution of estrogen (ER), progesterone (PR) and androgen receptors (AR) in the vagina and to determine if estradiol treatment ameliorates these changes. Eight-week-old female diabetic (db/db) mice (strain BKS.Cg-m+/+ Lepr^db/J) were divided into two subgroups: untreated diabetic and diabetic animals treated with pellets containing estradiol. Control normoglycemic littermates were subcutaneously implanted with pellets devoid of estradiol. At 16 weeks of age, animals were sacrificed, vaginal tissues excised and analyzed by Western blot and immunohistochemical methods. Diabetes produced marked reductions in protein expression of ER, PR, and AR. Diabetes also resulted in marked differences in the distribution, staining intensity and proportion of immunoreactive cells containing these steroid receptors in the epithelium, lamina propria and muscularis. Treatment of diabetic animals with estradiol restored receptor protein expression and distribution similar to those levels observed in control animals. This study demonstrates that type 2 diabetes markedly reduces steroid receptor protein expression and distribution in the vagina. Estradiol treatment of diabetic animals ameliorates these diabetes-induced changes.     

Effect of Ras Inhibition in Hematopoiesis and BCR/ABL Leukemogenesis

Introduction

Breast cancer is the most common female cancer and the second most common cause of female cancer-related deaths in the United States. World-wide, more than one million women will be diagnosed with breast cancer annually. In 2007, more than 175,000 women were diagnosed with breast cancer in the United States. However, deaths due to breast cancer have decreased in the recent years in part because of improved screening techniques, surgical interventions, understanding of the pathogenesis of the disease, and utilization of traditional chemotherapies in a more efficacious manner. One of the more exciting areas of improvement in the treatment of breast cancer is the entrance of novel therapies now available to oncologists. In the field of cancer therapeutics, the area of targeted and biologic therapies has been progressing at a rapid rate, particularly in the treatment of breast cancer. Since the advent of imatinib for the successful treatment of chronic myelogenous leukemia in the 2001, clinicians have been searching for comparable therapies that could be as efficacious and as tolerable. In order for targeted therapies to be effective, the agent must be able to inhibit critical regulatory pathways which promote tumor cell growth and proliferation. The targets must be identifiable, quantifiable and capable of being interrupted. In the field of breast cancer, two advances in targeted therapy have led to great strides in the understanding and treatment of breast cancer, namely hormonal therapy for estrogen positive receptor breast cancer and antibodies directed towards the inhibition of human epidermal growth factor receptor (HER)2. These advances have revolutionized the understanding and the treatment strategies for breast cancer. Building upon these successes, a host of novel agents are currently being investigated and used in clinical trials that will hopefully prove to be as fruitful. This review will focus on novel therapies in the field of breast cancer with a focus on metastatic breast cancer (MBC) and updates from the recent annual ASCO meeting and contains a summary of the results.     

Hormone response element binding proteins: novel regulators of vitamin D and estrogen signaling

Insights from vitamin D-resistant New World primates and their human homologues as models of natural and pathological insensitivity to sterol/steroid action have uncovered a family of novel intracellular vitamin D and estrogen regulatory proteins involved in hormone action. The proteins, known as “vitamin D or estrogen response element-binding proteins”, behave as potent cis-acting, transdominant regulators to inhibit steroid receptor binding to DNA response elements and is responsible for vitamin D and estrogen resistances. This set of interactors belongs to the heterogeneous nuclear ribonucleoprotein (hnRNP) family of previously known pre-mRNA-interacting proteins. This review provides new insights into the mechanism by which these novel regulators of signaling and metabolism can act to regulate responses to vitamin D and estrogen. In addition the review also describes other molecules that are known to influence nuclear receptor signaling through interaction with hormone response elements.     

Cognition, mood disorders, and sex hormones

Macaques (Macaca spp.) are useful models to evaluate effects of ovarian sex steroids and selective estrogen receptor modulators (SERMs) on mood and cognitive function due to similarities to women in their reproductive and central nervous systems. The results of nonhuman primate studies support the hypothesis that estrogen mediates specific aspects of attention and memory, yet much work is needed to understand which cognitive processes are affected, whether natural versus surgical menopause effects are different, and the interaction of age and ovarian senescence on cognitive function. This knowledge is necessary to determine whether to support the cognitive function of women in the menopausal phase of life and, if so, to determine efficacious therapeutic interventions. Mood disorders are prevalent in women and are associated with reproductive function in women and macaques. Exogenous steroid therapies, including oral contraceptives and postmenopausal hormone replacement therapies, have behavioral effects in women and appear to affect the behavior and underlying neural substrates of monkeys. Additional research is necessary to confirm and extend these observations. Ovarian steroids have multiple effects on serotonin synthesis, reuptake, and degradation, on neural activity that drives serotonin release, and on receptor activation in primates. This system modulates cognitive function and mood and is the target of a broad class of antidepressant therapies. Understanding the effects of ovarian steroids on the neural serotonergic system is necessary to understand depression in women. These studies are best carried out in primate models, which are more similar to humans in neural serotonergic function than other animal models.     

Tissue selectivity of ospemifene: Pharmacologic profile and clinical implications

The multifactorial consequences of menopausal estrogen deficiency affect numerous tissues throughout the body. Supplemental hormonal therapies carry the burden of a risk/benefit ratio that must be highly individualized. Selective estrogen receptor modulators (SERMs) are estrogen receptor (ER) agonist/antagonists designed to induce benefits comparable with estrogen while minimizing adverse effects. Here, we review the estrogen agonist/antagonist profile of ospemifene, a novel triphenylethylene derivative recently approved to treat dyspareunia, a symptom of vulvar and vaginal atrophy (VVA) due to menopause, both preclinically and clinically. Ospemifene binds ERα and ERβ with approximately equal affinities. In preclinical models, ospemifene increased vaginal and uterine epithelial thickness and mucification to the same extent as estrogen. Ospemifene did not induce endometrial hyperplasia in animal models; there also was no stimulatory effect on endometrial cells. In rat and human mammary cells in vitro, ospemifene evokes a dose-dependent inhibition on estrogen-induced cell responses and cell proliferation, supporting an antiestrogenic effect in breast. In contrast, ospemifene has an estrogenic effect on bone, as seen by improved bone mineral density, strength, mass, and histomorphometry in preclinical models, consistent with improvements in markers of bone resorption and formation in postmenopausal women. Based on the preclinical evidence, ospemifene has beneficial estrogen-like effects on the vaginal epithelium, preliminary evidence to support a neutral endometrial profile, antiproliferative effects in breast, and estrogenic effects in bone. Taken together, especially regarding estrogen-like effects on the vaginal epithelium, ospemifene presents a profile of tissue-specific effects that appear novel among available SERMs and well-suited for the treatment of VVA.     

Flow cytometric analysis of human breast tumors and assessment of in vitro chemosensitivity by clonogenic assays

We report a double-agar clonogenic system adapted to human breast cancer. We optimized the conditions for cell growth and clonogenicity with respect to hormones (insulin, estradiol, progesterone) and components of the extracellular matrix (collagen, laminin and fibronectin). Using our experimental improvements, 67% of the breast tumor samples received were grown successfully. Tests on 21 tumors with three agents: Doxorubicin, Methotrexate and 5-Fluorouracil permit objective discrimination of the in vitro pharmacosensitivity of human breast tumors. Flow cytometric analysis reveal that 64% of the tumors were diploid and 36% were aneuploid. The aneuploid tumors grew better in the double agar layer system used for the clonogenic assay. The diploid tumors were especially rich in estrogen (ER⁺) and progesterone (PR⁺) receptors whereas the aneuploid tumors were mostly estrogen and progesterone receptors negative (ER^–/PR^–). Finally, we noted no difference in drug responsiveness depending on the tumor ploidy and steroid receptor content.Abbreviations DCC dextran coated charcoal - DI DNA index - DXB Doxorubicin - ECM extracellular matrix component - ER estrogen receptors - FCM flow cytometry - 5-FU 5-Fluorouracil - HTSCA human tumor stem cell assay - MTX Methotrexate - PBC primary breast carcinoma - PI proliferative index - PR progesterone receptors - SPF S phase fraction     

Routine assessment of hormonal receptor and her-2/neu status underscores the need for more therapeutic targets in Kenyan women with breast cancer

OBJECTIVE: To report the expression of estrogen receptors, progesterone receptors and human epidermal growth factor receptor (Her-2/neu) in 158 Kenyan women with breast cancer and correlation with other prognostic indicators in this high-risk group. This study stressed the importance of routine assessment of the steroid receptors and Her-2/neu as a mode of therapeutic selection of patients for antihormonal or targeting monoclonal antibody (Herceptin) therapy, directed at the juxtamembrane domain of Her-2/neu protein in the developing countries such as Kenya. STUDY DESIGN: The study population consisted of 158 female patients with histologically confirmed breast carcinoma seen at the pathology department of The Nairobi Hospital. An immunohistochemical (IHC) study of ER, PR and Her-2/neu was conducted, followed by fluorescent in situ hybridization (FISH) validation for Her-2/neu gene amplification in cases initially scored as positive 2+ with IHC. Mastectomy samples registered at the pathology department of The Nairobi Hospital were used for this study. The study was approved by the institution's ethical review committee and informed consent obtainedfrom the concerned patients. RESULTS: In the studied cohort, positivity for both hormonal receptors and Her-2/neu was noted in 10 (6.33%) cases and negativity in 44 (27.85%) cases. Conversely, Her-2/neu negativity was noted in 32 (20.25%) cases with both steroid receptors positive and Her-2/neu positivity with both steroid receptors negative in 20 (12.66%) cases. Overall, no predictive factor was found in the Her-2/neu amplified 31/153 (20.26%) cases completely assessed with IHC and FISH. Grade III invasive ductal carcinomas, however, had a high prevalence of Her-2/neu overexpression. Association of both menopausal status (p = 0.044) and progesterone receptor status (p = 0.004) with high grade tumors were found to be statistically significant at 95% CI (p < 0.5). Consistent with other studies, Her-2/neu overexpression in this cohort was 20.26%. CONCLUSION: Her-2/neu positivity may activate ER expression through signaling kinases, and the combined target of mitogenic estrogen plus the monoclonal antibody therapy against Her-2/neu-overexpressing tumors expand chances of survival for patients in developing countries such as Kenya. The cost factor for these tests, selection for appropriate combined therapies and lack of awareness were noted as limiting factors for access to basic health care service and resulted in advanced tumor grade at time of patient presentation.     

Estrogen induces death of tamoxifen-resistant MCF-7 cells: contrasting effect of the estrogen receptor downregulator fulvestrant

A common problem in breast cancer therapy is resistance to the antiestrogen tamoxifen. However, tamoxifen-resistant breast tumors can still respond to other hormonal therapies. In animal models of tamoxifen-resistant breast cancer cells, physiological levels of estrogen can induce tumor regression. Recently, the estrogen receptor downregulator fulvestrant was shown to promote tumor growth of tamoxifen-resistant cells when added in combination with physiological levels of estrogen. Here, we show, using a cell culture model, that continuous exposure of tamoxifen-resistant cells to physiological levels of estrogen leads to cell death. Addition of the estrogen receptor downregulator fulvestrant prevents estrogen-induced death in a dose-dependent manner. Our data indicate that endogenous levels of estrogen affect the response of tamoxifen-resistant cells to fulvestrant. These results suggest that failure of fulvestrant to inhibit tumor growth in some tamoxifen-resistant patients may be due to endogenous estrogen levels. Moreover, these studies support short-term treatment with estrogen as a second-line hormonal therapy for tamoxifen-resistant breast cancer.