High Doses Of Estradiol Research Articles

Reduction of Kiss1 expression in the anteroventral periventricular nucleus is associated with atrazine-induced attenuation of the luteinizing hormone surge in female rats.

Atrazine, a commonly used herbicide, suppresses the luteinizing hormone (LH) surge in female rats, although the underlying mechanism remains unclear. Kisspeptin, encoded by the Kiss1 gene, is a hypothalamic peptide that controls gonadotropin-releasing hormone (GnRH) release from the GnRH neurons. Kisspeptin neurons in the anteroventral periventricular nucleus (AVPV) are involved in regulating pre-ovulatory GnRH and LH surge. To clarify the effect of atrazine on the LH surge in female rats, we investigated its effects on hypothalamic GnRH and kisspeptin. Ovariectomized female rats in a high-dose estradiol supplementation model were orally administered vehicle or 100 mg/kg of atrazine once daily for 5 days. This attenuated the LH surge but did not affect baseline LH levels, with no difference in hypothalamic GnRH levels between the vehicle-treated and atrazine-treated animals. After the fifth treatment, subcutaneous administration of kisspeptin (at 0, 0.1, 1, and 10 nmol/kg) induced a dose-dependent LH release almost equivalent in the vehicle- and atrazine-treated animals, suggesting that GnRH neurons maintain normal responsiveness to kisspeptin. However, Kiss1 mRNA expression levels in the AVPV were significantly reduced in the atrazine-treated animals. Given the normal response of GnRH neurons to exogenously administered kisspeptin, the suppressive effect of atrazine may be explained by suppression of Kiss1 expression in the AVPV leading to the attenuation of kisspeptin release from kisspeptin neurons in the AVPV. Further studies are warranted to elucidate more precisely the mechanism of atrazine's involvement in the suppression of Kiss1 mRNA expression in the AVPV.

Impact of Perfluorooctane Sulfonate on Reproductive Ability of Female Mice through Suppression of Estrogen Receptor α-Activated Kisspeptin Neurons.

Perfluorooctane sulfonate (PFOS) is used extensively in industrial and household applications. High exposure to PFOS has been associated with increased odds of irregular and long menstrual cycles in women. However, the underlying mechanisms remain to be elucidated. Herein, we show that adult female mice appeared prolongation of diestrus and reduction of corpora luteum within a week of oral administration of PFOS (10 mg/kg), which are associated with decreases in the levels of serum progesterone, LH and hypothalamic GnRH. The number of AVPV-kisspeptin neurons and the AVPV-kisspeptin expression were increased in proestrus mice or OVX-mice treated with high-dose estradiol benzoate (0.05 mg/kg), which were suppressed by the administration of PFOS. The administration of PFOS or GPR54 antagonist P234 prevented the generation of LH-surge in OVX-mice treated with high-dose E2. In hypothalamic slices incubated in 100 nM E2 for 4 h, the AVPV-kisspeptin expression was significantly enhanced, which was inhibited by PFOS in a dose-dependent manner or estrogen receptor α (ERα) antagonist MPP, but not ERβ antagonist PHTPP. The incubation of ERα agonist PPT rather than ERβ agonist DPN could increase the level of AVPV-kisspeptin expression, which was sensitive to the treatment with PFOS. The administration of GPR54 agonist kisspeptin-10 in PFOS-mice could correct the prolongation of diestrus and reduction of corpora luteum, and recover the LH-surge and the levels of LH and GnRH. The results indicate that exposure to PFOS suppressed ERα-induced activation of AVPV-kisspeptin neurons leads to diestrus prolongation and ovulation reduction.

Effects of systemic estradiol on fear extinction in female rats are dependent on interactions between dose, estrous phase, and endogenous estradiol levels

Administering estradiol to females during periods of low endogenous estradiol enhances their ability to extinguish fear, the laboratory basis of exposure therapy for anxiety disorders. It has therefore been proposed that estradiol could be a useful adjunct to enhance exposure therapy outcomes. The present study aimed to clarify the boundary conditions under which estradiol could be used for this purpose, by assessing whether the impact of estradiol, administered systemically prior to extinction training, differs depending on dose and estrous phase in adult female rats. Results demonstrated that in rats extinguished during metestrus (naturally low estradiol), a low dose of estradiol reduced freezing during extinction training and augmented extinction recall the following day, whereas a high dose of estradiol had no effect on either extinction training or recall. In rats extinguished during proestrus (naturally high estradiol), a high dose of estradiol impaired extinction recall, whereas a low dose of estradiol had no effect, or impairing effects, on extinction recall in different experiments. A subsequent analysis revealed that estradiol-treated proestrus rats that exhibited impaired extinction recall had significantly higher pre-treatment serum estradiol levels than those that exhibited good extinction recall. Together, these results indicate that systemically administered estradiol interacts with endogenous estradiol to produce an inverted U shaped dose effect on fear extinction, where low and high estradiol levels lead to poor extinction recall, and moderate estradiol levels lead to good extinction recall. These results highlight potential limitations to the use of estradiol as an adjunct to exposure therapy in clinical settings.

Retraction Note to: Extranuclear ER\u03b1 is associated with regression of T47D PKC\u03b1-overexpressing, tamoxifen-resistant breast cancer

Prior to the introduction of tamoxifen, high dose estradiol was used to treat breast cancer patients with similar efficacy as tamoxifen, albeit with some undesirable side effects. There is renewed interest to utilize estradiol to treat endocrine resistant breast cancers, especially since findings from several preclinical models and clinical trials indicate that estradiol may be a rational second-line therapy in patients exhibiting resistance to tamoxifen and/or aromatase inhibitors. We and others reported that breast cancer patients bearing protein kinase C alpha (PKCα)- expressing tumors exhibit endocrine resistance and tumor aggressiveness. Our T47D:A18/PKCα preclinical model is tamoxifen-resistant, hormone-independent, yet is inhibited by 17β-estradiol (E2) in vivo. We previously reported that E2-induced T47D:A18/PKCα tumor regression requires extranuclear ERα and interaction with the extracellular matrix. T47D:A18/PKCα cells were grown in vitro using two-dimensional (2D) cell culture, three-dimensional (3D) Matrigel and in vivo by establishing xenografts in athymic mice. Immunofluoresence confocal microscopy and co-localization were applied to determine estrogen receptor alpha (ERα) subcellular localization. Co-immunoprecipitation and western blot were used to examine interaction of ERα with caveolin-1. We report that although T47D:A18/PKCα cells are cross-resistant to raloxifene in cell culture and in Matrigel, raloxifene induces regression of tamoxifen-resistant tumors. ERα rapidly translocates to extranuclear sites during T47D:A18/PKCα tumor regression in response to both raloxifene and E2, whereas ERα is primarily localized in the nucleus in proliferating tumors. E2 treatment induced complete tumor regression whereas cessation of raloxifene treatment resulted in tumor regrowth accompanied by re-localization of ERα to the nucleus. T47D:A18/neo tumors that do not overexpress PKCα maintain ERα in the nucleus during tamoxifen-mediated regression. An association between ERα and caveolin-1 increases in tumors regressing in response to E2. Extranuclear ERα plays a role in the regression of PKCα-overexpressing tamoxifen-resistant tumors. These studies underline the unique role of extranuclear ERα in E2- and raloxifene-induced tumor regression that may have implications for treatment of endocrine-resistant PKCα-expressing tumors encountered in the clinic.

Estrogen Status Gates Effects of Kappa-Opioid Receptor on Temporomandibular Joint-Responsive Neurons at the Spinomedullary Junction in Female Rats.

To determine whether estrogen status alters κ-opioid inhibition of nociceptive processing by affecting temporomandibular joint (TMJ) input to neurons in the trigeminal subnucleus caudalis [Vc]/C1-2 region at the spinomedullary junction in female rats. TMJ-responsive neurons were recorded in laminae I-II of the Vc/C1-2 region at the spinomedullary junction of ovariectomized female rats treated for 2 days with low-dose estradiol (LE group; 2 mg/day) or high-dose estradiol (HE group; 20 mg/day). Under isoflurane anesthesia, TMJ neurons were activated by adenosine triphosphate (ATP; 1 mM, 20 μl), which was injected into the joint space before and after cumulative doses of a κ-opioid receptor (KOR) agonist (U50488) given systemically (0.03, 0.3, and 3 mg/kg, intravenously) or by local application to the dorsal surface of the Vc/C1-2 region (1 and 10 nmol/30 μl). Analysis of variance and Newman-Keuls test were performed to compare the data. Systemic U50488 caused a dose-related inhibition of ATP-evoked neuronal activity in HE rats and reduced the size of the neuronal cutaneous receptive field (RF), while effects in LE rats were not significant. Systemic U50488 reduced the spontaneous activity of TMJ-responsive neurons to similar levels in LE and HE groups. Locally applied U50488 inhibited ATP-evoked neuronal activity in HE rats, but not in LE rats. Systemic and local administration of the KOR antagonist nor-binaltorphinine (nor-BNI) partially reversed the decrease in Rmag induced by U50488, but had no effect on neurons from LE rats. These results indicate that KOR-dependent effects on TMJ-responsive neurons in the superficial laminae of the Vc/C1-2 region in female rats are differentially modified by high and low estrogen status. The site of action for estrogen-induced modulation of TMJ neuronal activity by KOR likely includes second-order neurons in the Vc/C1-2 region.

Efficacy, maximal tolerated dose, and toxicokinetics of TTC-352 in rats and dogs, a partial ER agonist for metastatic ER+ breast cancer.

e14090 Background: The high prevalence of treatment resistance for estrogen receptor positive (ER+) breast cancer cause more deaths than all other breast cancers, despite the availability of endocrine therapeutics including selective ER downregulators (SERDs). Before tamoxifen, high dose estradiol (E2) delivered clinical efficacy slightly superior to tamoxifen. While recent clinical trials have shown efficacy of low dose E2, unacceptable side effects including vaginal bleeding, endometrial hypertrophy, and the negative perceptions of both patients and physicians limit acceptance. In preclinical models, tamoxifen resistant (TR) breast cancer is sensitive to treatment by E2. TTC-352 is an orally bioavailable selective human ERα partial agonist (ShERPA) designed to mimic E2 in causing tumor regression of TR breast cancer xenografts without uterine proliferation caused by E2 and tamoxifen. Methods: The published preclinical efficacy studies were extended to multiple TR ER+ cell lines in 3D spheroid cell cultures comparing to E2 and the SERD GDC0810. The maximum tolerated dose (MTD) was determined in Sprague-Dawley rats at: 200, 300, 600, 1000 and 2000 mg/kg by oral gavage (N = 3). Animals were observed for 7 days prior to necropsy. A single MTD range finding study was performed in dogs at: 50, 100, and 200 mg/kg TTC-352, administered orally in gelatin capsules. Based on these results, a 7-day repeated dose studies were completed at 30, 100, 300 and 1000 mg/kg/day in rats (3/sex/dose) and 15, 75, and 150 mg/kg (2/sex/dose) in dogs. Results: Growth of three TR ER+ breast cancer cell cultures in 3D was inhibited by TTC-352, which mimicked E2 and was equivalent to GDC0810. In female rats MTD was 1000 mg/kg and TTC-352 was well tolerated following a single oral dose of 200, 300, 600 mg/kg. The oral administration of TTC-352 at doses of 30, 100, and 300 mg/kg/day in rats (both sexes) and 15, 75 or 150 mg/kg/day in dogs (both sexes) for seven days was generally well tolerated. Conclusions: TTC-352 has demonstrated efficacy in preclinical models of TR breast cancer. Completed rat and dog studies indicated favorable tolerability and rapid absorption up to 300 mg/kg/day in rats and 150 mg/kg/day in dogs.

Evaluating the role of astrocytes on β-estradiol effect on seizures of Pilocarpine epileptic model

Epilepsy with periodic and unpredictable seizures is associated with hippocampal glutamate toxicity and tissue reorganization. Astrocytes play an important role in mediating the neuroprotective effects of estradiol and reducing seizure severity. Accordingly, the protective effects of low and high doses of estradiol on behavioral, astrocytic involvement and neuronal survival aspects of Pilocarpine-induced epilepsy were investigated. Lithium- Pilocarpine (30mg/kg) model was used to provoke epilepsy. Βeta-estradiol (2,40μg/µl) was injected subcutaneously from 48 before to 48h after seizure induction. Behavioral convulsions were then monitored and recorded on the day of induction. Four weeks later, glutamine synthetase (GS) activity and the astrocyte transporter GLT-1 expression of the hippocampus were measured. Moreover, hippocampal glutamate and GABA were evaluated to study excitability changes. Finally, neuronal counting in the hippocampus was also performed using Nissl staining. The latency for generalized clonic (GC) convulsions significantly increased while the rate of GC and death significantly reduced due to β-estradiol treatment. GS activity and GLT-1 expression increased in the groups receiving the high dose of β-estradiol and Pilocarpine. Furthermore, the amount of both GABA and glutamate content decreased due to high dose of estradiol, while only GABA increased in Pilocarpine treated rats. Finally, administration of β-estradiol with low and high doses increased and improved the density of nerve cells. It is concluded that chronic administration β-estradiol has anticonvulsant and neuroprotective properties which are plausibly linked to astrocytic activity.

Targeting Estrogen Receptor Beta in a Phase 2 Study of High-Dose Estradiol in Metastatic Triple-Negative Breast Cancer: A Wisconsin Oncology Network Study

BackgroundEstrogen receptor beta (ERβ) is expressed by 50% to 80% of triple-negative breast cancers (TNBC). Agonism of ERβ has antiproliferative effects in TNBC cells expressing ERβ. This phase 2 study evaluated single-agent high-dose estradiol in patients with advanced TNBC. Patients and MethodsAdult women with measurable advanced TNBC were treated with estradiol 10 mg oral 3 times daily provided continuously for 28-day cycles. A Simon optimal 2-stage design was used. The primary end point was objective response (OR). Secondary end points included progression-free survival (PFS), clinical benefit (CB), and safety. OR, CB, and PFS by ERβ status were also examined. ResultsSeventeen evaluable women were enrolled. Median age was 58 years (range, 34-90 years); the median number of prior systemic therapies was 2 (range, 0-6). One patient had a confirmed partial response (OR rate, 5.9%) and remained on the study for > 24 weeks. Three patients had stable disease, with one lasting more than 16 weeks. ERβ expression was detected in 77% (13 patients). The CB rate at 16 weeks was 15% (2 of 13) in ERβ-positive patients and 0% (0 of 4) in ERβ-negative patients (P = 1). PFS was poor (median, 1.9 months) and not statistically significantly different between ERβ-positive versus -negative patients. No new adverse events from estradiol were identified. The study closed after the first stage as a result of limited responses in these unselected patients. ConclusionIn unselected TNBC, high-dose estradiol has limited efficacy. However, further evaluation of ERβ selective agonists in TNBC selected by ERβ expression may be warranted.

Lectin-mediated delivery of α-L-iduronidase: A novel approach for MPS I enzyme replacement therapy

Epilepsy with periodic and unpredictable seizures is associated with hippocampal glutamate toxicity and tissue reorganization. Astrocytes play an important role in mediating the neuroprotective effects of estradiol and reducing seizure severity. Accordingly, the protective effects of low and high doses of estradiol on behavioral, astrocytic involvement and neuronal survival aspects of Pilocarpine-induced epilepsy were investigated. Lithium- Pilocarpine (30 mg/kg) model was used to provoke epilepsy. Βeta-estradiol (2,40 μg/µl) was injected subcutaneously from 48 before to 48 h after seizure induction. Behavioral convulsions were then monitored and recorded on the day of induction. Four weeks later, glutamine synthetase (GS) activity and the astrocyte transporter GLT-1 expression of the hippocampus were measured. Moreover, hippocampal glutamate and GABA were evaluated to study excitability changes. Finally, neuronal counting in the hippocampus was also performed using Nissl staining. The latency for generalized clonic (GC) convulsions significantly increased while the rate of GC and death significantly reduced due to β-estradiol treatment. GS activity and GLT-1 expression increased in the groups receiving the high dose of β-estradiol and Pilocarpine. Furthermore, the amount of both GABA and glutamate content decreased due to high dose of estradiol, while only GABA increased in Pilocarpine treated rats. Finally, administration of β-estradiol with low and high doses increased and improved the density of nerve cells. It is concluded that chronic administration β-estradiol has anticonvulsant and neuroprotective properties which are plausibly linked to astrocytic activity.

Assessment of estradiol-induced gene regulation and proliferation in an immortalized mouse immature Sertoli cell line

AimsThe number of Sertoli cells during proliferative phase determines the fate of the germ cells in male reproductive system. A well-characterized cell line may help in better understanding of Sertoli cell biology. Hence, the present study assessed estradiol signaling in a mouse immature Sertoli cell line (MSC-1) as an alternative model in place of primary culture of Sertoli cells. Main methodsIn this study, we used MSC-1 cell line, derived from 10-day old mice. The cell cycle parameters were assessed, and the expression and regulation of Sertoli cell-specific secretory genes (ABP; androgen-binding protein) and tight junction genes (claudin-5, occludin, and vimentin) in response to estradiol was studied. Key findingsThe results obtained suggested the presence of both estrogen receptors (ERα and ERβ) in MSC-1 cells. In vitro scratch assay and cell-cycle analysis suggested the proliferative effects of estradiol in both time- and dose-dependent manner. The gene expression profiles of ABP, claudin-5, and occludin showed biphasic regulation at low and high doses of estradiol. Analysis of signaling pathways suggested the activation of extracellular signal-regulated kinase (ERK) pathway with significantly increased pERK/ERK ratio (p<0.05). The results also suggested down regulation in the expression of mir-17 family members (mir-17, mir-20b, and mir-106a) (p<0.05). SignificanceConsidering the limited number of Sertoli cell lines and long-term survival inability of primary culture of Sertoli cells, MSC-1 cells could be a potential cell line for understanding the mechanisms of various cellular events in Sertoli cells.

Effect of estrodiol on leptin receptors expression in regulating fat distribution and adipocyte genesis

Objective: To examine how estradiol influence fat distribution and adipocyte genesis through regulating leptin function.Design and methods: We randomized female SD rats into ovariectomy and sham group. After 14 weeks, we examined leptin receptor expression in perigenital, mesenteric and subcutaneous adipose tissues and leptin in serum. We further introduced mesenchymal stem cells (MSCs) to figure out the effect of 17-β estradiol on expression of leptin receptors (OBRb and OBRa), leptin and PPARγ2.Results: Weight and Lee’s index was both significantly higher in ovariectomized group than sham group (p < 0.001). The interaction of serum E2 and leptin was negatively correlated with body weight. Expression of leptin receptor protein was extremely upregulated in ovariectomized group in perigenital and mesenteric fat. Both estradiol and leptin upregulated mRNA expression of OBRb, and created a dose-dependent decreasing manner in MSCs. Higher doses of estradiol (10−7–10−5 M) inhibited adipogenic markers mRNA expression of leptin and PPARγ2, but low doses promoted leptin expression.Conclusions: In ovariectomized rats, low level of serum estradiol may change body composition and promote adipocyte genesis through affecting leptin function.

Hormonal stimulation and paternal experience influence responsiveness to infant distress vocalizations by adult male common marmosets, Callithrix jacchus

Parental experience and hormones play a large role in the common marmoset (Callithrix jacchus) father's care of their offspring. We tested the effect of exogenous estradiol or testosterone on the responsiveness of common marmosets to respond to infant distress vocalizations and whether males who haven't become fathers yet (paired males) would have increased responsiveness to infant distress calls with either steroid or whether parental experience is the most important component for the onset of paternal care. Sixteen male marmosets (8 fathers, 8 paired males) received a vehicle, low dose or high dose of estradiol and additional 16 males were tested with testosterone at three doses for their response either to a vocal control or a recording of an infant distress call for 10min. Without steroid stimulation fathers were significantly more likely to respond to the infant distress stimulus than paired males. Low dose estradiol stimulation resulted in a significant increase in fathers' behavioral response towards the infant distress stimulus but not in paired males. Fathers also showed a significant increase in infant responsiveness from the vehicle dose to the estradiol low dose treatment, but not to the estradiol high dose treatment. Testosterone treatment did not show significant differences between infant responsiveness at either dose and between fathers and paired males. We suggest that neither steroid is involved in the onset of paternal care behaviors in the marmoset but that estradiol may be involved in facilitating paternal motivation in experienced fathers.

Food restriction-induced augmentation of heroin seeking in female rats: manipulations of ovarian hormones.

Food restriction augments heroin seeking in chronically food-restricted male rats under withdrawal, an effect not yet examined in female rats. Importantly, women and female rats possess an increased vulnerability to drugs of abuse, which may be mediated by fluctuations in ovarian hormones. We investigated the role of estradiol and progesterone in augmented heroin seeking in chronically food-restricted female rats, under withdrawal. Female rats self-administered heroin for 10-12 days and were then allowed unrestricted (sated) or restricted access to food (FDR; ∼10 % reduction in body weight) for 14 days. On day 14, rats underwent a heroin-seeking test. Exp. 1: Rats underwent ovariectomy or sham surgery and were treated with a low dose of estradiol (5.0 % in cholesterol; subcutaneous capsule). Exp. 2: Rats underwent ovariectomy and were administered with a high dose of estradiol (0.5 mg/kg; subcutaneous) for 8 days before testing. Exp. 3: Progesterone injections (2.0 mg/kg; subcutaneous) were administered 24 h and 2 h before testing. Food restriction resulted in augmented heroin seeking, compared to sated controls. While ovariectomy had no effect, estradiol replacement attenuated the food restriction effect. Injections of progesterone had no effect on heroin seeking in either the sated or FDR groups. The effect of food restriction on heroin seeking in female rats under withdrawal is as robust as previously found in males. Interestingly, estradiol replacement, but not progesterone, attenuates the food restriction effect in the ovariectomized rats, possibly due to its anorexic properties.

Changes in estrogen receptor signaling alters the timekeeping system in male mice

Circadian rhythms are modulated by steroid hormones; however, the mechanisms of this action are not fully understood, particularly in males. In females estradiol regulates activity level, pattern of expression, and free running period (tau). We tested the hypothesis that activity level and distribution in male mice includes both classical and “non-classical” actions of estrogens at the estrogen receptor subtype 1 (ESR1). We used transgenic mice with mutations in their estrogen response pathways: ESR1 knock-out (ERKO) mice lack the ability to respond to estrogens via ESR1. “Non-classical” estrogen receptor knock-in (NERKI) mice have an inserted ESR1 receptor with a mutation in the estrogen-response-element binding domain, allowing activation via non-genomic and second messenger pathways. Gonadectomized male NERKI, ERKO, and wildtype (WT) littermates were given oil, or low or high dose estradiol and daily activity parameters were quantified. Estradiol shortened the ratio of activity in the light relative to dark (LD ratio), shortened tau, advanced the time of activity onset, and altered responsiveness to light cues administered in the late subjective night, suggesting modulation by an ESR1-independent mechanism. Estradiol treatment in NERKI but not WT males altered the timing of activity onset, LD ratio, and the behavioral response to light cues. These results may represent disruptions in the balance of genomic/nongenomic or ESR1/ESR2 signaling pathways. We also found a significant genotype effect on total activity, LD ratio, tau, and activity duration. These data provide new information about the role of ESR1-dependent and independent signaling pathways on the timekeeping system in male mice.

Abstract PD6-5: Profiling of ESR1-mutated metastatic breast cancers by FoundationOne® allows a broad genomic understanding for potential clinical implications

Abstract Background: Estrogen receptor (ER) inhibition is an important treatment option for advanced breast cancer (BC) pts. Recent studies describe recurring somatic mutations in codons 537 and 538 within the ligand-binding domain (LBD) of ∼ 20% of ER+ metastatic disease, but not treatment-naïve ER+ cancer or ER-negative disease, that render the ER constitutively active and confer partial resistance to endocrine agents. Few studies have described the clinicopathologic and genomic covariates that accompany the ESR1 genomic alterations (GAs). Correlating genomic events as captured by FoundationOne in pts with ESR1-mutated BCs with clinical history may provide clinical insight into these alterations. Methods: Hybridization capture of 3769 exons of 236 cancer-related genes and 47 introns from 19 genes that are frequently rearranged in cancer were fully sequenced to high, uniform coverage using FoundationOne (Foundation Medicine, Cambridge, MA). Results: 176 of the 2,208 (7.9%) BC pt cases harbored ESR1 alterations. 1127 short variants (SV) were detected in 176 patient samples for an average of 6.4 GA/sample. 16.5% SV (186/1127) were ESR1 GAs with an average of 1.0 ESR1 GAs/sample. ESR1 GAs consisted of base substitutions (77%) and amplifications (20%, median copy number 9X, range 6-28). Base substitutions occurred at codons 538 (71/145) and 537 (70/145) and at two other novel sites, 341 (2/145) and 563 (1/145). A patient-derived xenograft study suggested that tumors refractory to ER inhibitors and harboring ESR1 amplification could be responsive to higher doses of estradiol. (Li et al, Cell Reports, 2013). The most frequently co-occurring GAs were PIK3CA (37.5%), GATA3 (22.7%), TP53 (24.4%), MAP3K1 (10.2%) and CDH1 (7.9). Collection of data on tissue sites sequenced, treatments and outcomes on these pts with ESR1 GAs are ongoing. Of note, one pt had recurrent disease following adjuvant tamoxifen and letrozole. FoundationOne testing of her recurrent bony disease revealed an ESR1 Y537 alteration along with mutations in PIK3CA, APC, and RAD51. She has been on fulvestrant monotherapy with stable metastatic disease 13 + months. Conclusions: We found through FoundationOne testing that ESR1 GAs are reasonably common in advanced BCs, with base substitutions accounting for 77% of ESR1 GAs and amplifications for 20%. Identification and characterization of ESR1-mutated advanced BC pts by comprehensive genomic profiling capable of detecting both base substitutions and copy number changes may identify clinically relevant GAs. Clinical correlation is pending. Citation Format: Norma A Palma, Siraj Ali, Garrett Frampton, Kai Wang, Hannah Gilmore, Julio Peguero, Lyndsay N Harris, Massimo Cristofanilli, Juliann Chmielecki, Jeffrey S Ross, Deborah Morosini, Vincent A Miller, Phil J Stephens, Gary Palmer, Joyce O'Shaughnessy. Profiling of ESR1-mutated metastatic breast cancers by FoundationOne® allows a broad genomic understanding for potential clinical implications [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 PD6-5.

Local group I mGluR antagonists reduce TMJ-evoked activity of trigeminal subnucleus caudalis neurons in female rats

Group I metabotropic glutamate receptors (mGluR1 and mGluR5) are functionally linked to estrogen receptors and play a key role in the plasticity of central neurons. Estrogen status strongly influences sensory input from the temporomandibular joint (TMJ) to neurons at the spinomedullary (Vc/C1–2) region. This study tested the hypothesis that TMJ input to trigeminal subnucleus caudalis/upper cervical cord (Vc/C1–2) neurons involved group I mGluR activation and depended on estrogen status. TMJ-responsive neurons were recorded in superficial laminae at the Vc/C1–2 region in ovariectomized (OvX) female rats treated with low-dose estradiol (2μg/day, LE) or high-dose estradiol (20μg/day, HE) for 2days. TMJ-responsive units were activated by adenosine triphosphate (ATP, 1mM) injected into the joint space. Receptor antagonists selective for mGluR1 (CPCCOEt) or mGluR5 (MPEP) were applied topically to the Vc/C1–2 surface at the site of recording 10min prior to the intra-TMJ ATP stimulus. In HE rats, CPCCOEt (50 and 500μM) markedly reduced ATP-evoked unit activity. By contrast, in LE rats, a small but significant increase in neural activity was seen after 50μM CPCCOEt, while 500μM caused a large reduction in activity that was similar in magnitude as that seen in HE rats. Local application of MPEP produced a significant inhibition of TMJ-evoked unit activity independent of estrogen status. Neither mGluR1 nor mGluR5 antagonism altered the spontaneous activity of TMJ units in HE or LE rats. High-dose MPEP caused a small reduction in the size of the convergent cutaneous receptive field in HE rats, while CPCCOEt had no effect. These data suggest that group I mGluRs play a key role in sensory integration of TMJ nociceptive input to the Vc/C1–2 region and are largely independent of estrogen status.

Gonadal hormones modulate the responsiveness to local β-blocker-induced antinociception in the temporomandibular joint of male and female rats.

We have previously demonstrated that blockade of β-adrenoreceptors (β-AR) located in the temporomandibular joint (TMJ) of rats suppresses formalin-induced TMJ nociceptive behaviour in both male and female rats, but female rats are more responsive. In this study, we investigated whether gonadal hormones modulate the responsiveness to local β-blocker-induced antinociception in the TMJ of rats. Co-administration of each of the selective β1 (atenolol), β2 (ICI 118.551) and β3 (SR59230A)-AR antagonists with equi-nociceptive concentrations of formalin in the TMJ of intact, gonadectomized and hormone-treated gonadectomized male and female rats. Atenolol, ICI 118.551 and SR59230A significantly reduced formalin-induced TMJ nociception in a dose response fashion in all groups tested. However, a lower dose of each β-AR antagonist was sufficient to significantly reduce nociceptive responses in gonadectomized but not in intact and testosterone-treated gonadectomized male rats. In the female groups, a lower dose of β1 -AR antagonist was sufficient to significantly reduce nociceptive responses in gonadectomized but not in intact or gonadectomized rats treated with progesterone or a high dose of oestradiol; a lower dose of β2 -AR antagonist was sufficient to significantly reduce nociceptive responses in gonadectomized but not in intact and gonadectomized rats treated with low or high dose of oestradiol. Gonadal hormones may reduce the responsiveness to local β-blocker-induced antinociception in the TMJ of male and female rats. However, their effect depends upon their plasma level, the subtype of β-AR and the dose of β-blockers used.

Abstract 2109: All trans-retinoic acid (ATRA) induces redifferentiation of early transformed breast epithelial cells

Abstract Retinoids have been used as potential chemotherapeutic or chemopreventive agents because of their differentiative, anti-proliferative, pro-apoptotic, and anti-oxidant properties. The effects of retinoids are mediated by retinoic acid receptors (RARβ;,RARα,RARγ) and the retinoid X receptors (RXR) that bind the DNA as heterodimers. The objective of this project was to study the potential of retinoic acid to revert the neoplastic process using an in vitro model of breast cancer progression. This model was previously developed by treating the human normal breast epithelial MCF-10F cells with high dose of estradiol and consists of four cell lines which show a progressive neoplastic transformation: (a) the spontaneously immortalized cell line MCF-10F, which does not show any characteristic of invasiveness or tumor formation and therefore is considered to be a normal-like breast epithelial cell line; (b) the transformed trMCF cells; (c) the invasive bsMCF cells and; (d) the caMCF tumor cells. The bsMCF cells induced tumors when they were injected in the mammary fat pad of SCID mice. We previously showed that RARβ was unmethylated in MCF-10F and trMCF cells, but became hypermethylated in bsMCF and caMCF. We studied the effect of retinoic acid at the different stages in this model by cultivating the cells in 3D-cultures (collagen matrix). In collagen, the MCF-10F cells form tubules resembling the normal mammary gland. After treatment with estradiol, cells formed tubules and spherical masses which are indicative of transformation. Cells that only formed spherical masses in collagen were isolated (trMCF clone 11) and treated with different concentrations of ATRA (10-5M to 10-8M). We found that trMCF clone 11 cells showed a reduction of solid masses and increased number of tubules in collagen after being treated with 10-6M ATRA. A total of 43% of the structures were tubules in trMCF clone 11 cells treated with 10-6M ATRA. Gene expression studies showed that 207 genes up-regulated in transformed trMCF clone 11 cells were down-regulated after ATRA treatment to levels comparable to those found in the normal breast epithelial cells MCF-10F. Furthermore, 236 genes that were down-regulated in trMCF clone 11 were up-regulated after ATRA treatment to similar levels shown in normal epithelial cells. These 443 genes defined a signature of the ATRA re-programming effect. Our results showed that 10-6M ATRA was able to re-differentiate transformed cells at early stages of the neoplastic process to a normal stage and antagonistically regulate breast cancer associated genes. The invasive and tumorigenic cells did not show any changes in morphology after treatment with ATRA alone or in combination with 5-aza-dC. These results suggest that ATRA could be used as chemopreventive agent to inhibit the progression of premalignant lesions of the breast. This work was supported by The Pennsylvania Breast Cancer Coalition and Friends for an Earlier Breast Cancer Test. Citation Format: Maria F. Arisi, Rebecca A. Starker, Sankar Addya, Yong Huang, Sandra V. Fernandez. All trans-retinoic acid (ATRA) induces redifferentiation of early transformed breast epithelial cells. [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 2109. doi:10.1158/1538-7445.AM2014-2109

Antidepressant May Have Role in Treating Menopause Symptoms

Back to table of contents Previous article Next article Clinical and Research NewsFull AccessAntidepressant May Have Role in Treating Menopause SymptomsJoan Arehart-TreichelJoan Arehart-TreichelSearch for more papers by this authorPublished Online:17 Jul 2014https://doi.org/10.1176/appi.pn.2014.7a6AbstractWhen menopausal women cannot or do not want to take estrogen to combat bothersome hot flashes and night sweats, antidepressants may serve as an effective alternative.Although estrogen has long been the gold standard for treating the hot flashes and night sweats of menopause, some women are unable or unwilling to use it because of associated risks. Consequently, SSRI or SNRI antidepressants—particularly the SNRI antidepressant venlafaxine—are often prescribed as a treatment for these vasomotor symptoms.Robert Kneschke/ShutterstockHowever, while SSRI and SNRI antidepressants have been found to be more effective than placebo in countering hot flashes and night sweats, their effectiveness compared with estrogen has not been studied. Hadine Joffe, M.D., an associate professor of psychiatry at Harvard Medical School, and her colleagues conducted a study to evaluate the two types of medications, using venlafaxine as the test antidepressant. As they reported May 26 in JAMA Internal Medicine, both estrogen and venlafaxine were significantly more effective than a placebo, but estrogen was slightly more effective than venlafaxine.A total of 339 perimenopausal and postmenopausal women with at least two bothersome vasomotor symptoms a day—on average, eight a day—took part in the study. The researchers excluded candidates who had experienced major depressive episodes within the previous year.The subjects were randomized to receive either low-dose oral estradiol (0.5 mg/d), low-dose venlafaxine extended release (75 mg/d), or a placebo for eight weeks. The researchers used low-dose rather than high-dose estradiol, they explained, “because of recommendations to use the lowest effective estrogen therapy dosage.”Throughout treatment, and on a daily basis, the subjects recorded in diaries how often they experienced vasomotor symptoms. By the end of treatment, the estradiol group was experiencing 3.9 vasomotor episodes a day on average, the venlafaxine group 4.4., and the placebo group 5.5. Estrogen reduced the frequency of vasomotor episodes by 53 percent, venlafaxine by 48 percent, and placebo by 29 percent. The differences between the estradiol group and the placebo group, as well as between the venlafaxine group and placebo group, were statistically significant. The differences between the venlafaxine group and the estradiol group were not.Tolerability of both estradiol and venlafaxine was high. Discontinuation because of adverse events was uncommon and did not differ significantly between treatments.The researchers also found that the treatment responses were not influenced by subjects’ mood symptoms, sleep quality, or race. A third of the subjects were African American.“It is nice to see this head-to-head study comparing estradiol and venlafaxine in the treatment of menopausal hot flashes,” Claudia Reardon, M.D., an assistant professor of psychiatry at the University of Wisconsin and an expert on women’s health issues, said in an interview with Psychiatric News. “While venlafaxine may not be quite as effective as estradiol in the treatment of these symptoms, it does appear to work notably better than placebo. Our women patients often talk to us as their mental health providers about their bothersome menopausal symptoms, and this study helps us to be able to discuss the likely relative efficacy of their options with them.”“The results of this carefully conducted study provide women and their physicians with critical data to guide treatment for vasomotor symptoms . . . , which affect the lives of the majority of midlife women,” Katherine Wisner, M.D., a professor of psychiatry and obstetrics and gynecology at Northwestern University, told Psychiatric News. “Although a number of studies have demonstrated the efficacy of venlafaxine and other serotonergic antidepressants for treating vasomotor symptoms, this study advances our knowledge by comparing estradiol and venlafaxine to placebo within the same study,” Laura Miller, M.D., medical director of women’s mental health at the Edward Hines Jr. VA Hospital in Illinois, said. “The findings confirm that both estradiol and venlafaxine reduce the frequency, severity, and interference of menopausal hot flashes. While supporting prior impressions that the impact of estradiol on vasomotor symptoms is greater than that of venlafaxine, it shows that the magnitude of difference is relatively small. Caveats are that this study is not a direct comparison between estradiol and venlafaxine (both were compared with placebo) and that medroxyprogesterone was not given during the study as it would be with clinical use of estrogen (it was given only after unblinding). [But] overall, the study adds valuable information about an alternative to estrogen for the treatment of bothersome menopausal vasomotor symptoms.”The study was funded by the National Institutes of Health. ■An abstract of “Low-Dose Estradiol and the Serotonin-Norepinephrine Reuptake Inhibitor Venlafaxine for Vasomotor Symptoms” can be accessed here. ISSUES NewArchived

All trans-retinoic acid (ATRA) induces re-differentiation of early transformed breast epithelial cells

Retinoids have been used as potential chemotherapeutic or chemopreventive agents because of their differentiative, anti-proliferative, pro-apoptotic and antioxidant properties. We investigated the effect of all trans-retinoic acid (ATRA) at different stages of the neoplastic transformation using an in vitro model of breast cancer progression. This model was previously developed by treating the MCF-10F human normal breast epithelial cells with high dose of estradiol and consists of four cell lines which show a progressive neoplastic transformation: MCF-10F, normal stage; trMCF, transformed MCF-10F; bsMCF, invasive stage; and caMCF, tumorigenic stage. In 3D cultures, MCF-10F cells form tubules resembling the structures in the normal mammary gland. After treatment with estradiol, these cells formed tubules and spherical masses which are indicative of transformation. Cells that only formed spherical masses in collagen were isolated (trMCF clone 11) and treated with ATRA. After treatment with 10 or 1 μM ATRA, the trMCF clone 11 cells showed tubules in collagen; 10 and 43% of the structures were tubules in cells treated with 10 and 1 μM ATRA, respectively. Gene expression studies showed that 207 genes upregulated in transformed trMCF clone 11 cells were downregulated after 1 μM ATRA treatment to levels comparable to those found in the normal breast epithelial cells MCF-10F. Furthermore, 236 genes that were downregulated in trMCF clone 11 were upregulated after 1 μM ATRA treatment to similar levels shown in normal epithelial cells. These 443 genes defined a signature of the ATRA re-programming effect. Our results showed that 1 μM ATRA was able to re-differentiate transformed cells at early stages of the neoplastic process and antagonistically regulate breast cancer associated genes. The invasive and tumorigenic cells did not show any changes in morphology after ATRA treatment. These results suggest that ATRA could be used as a chemopreventive agent to inhibit the progression of premalignant lesions of the breast.