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1.
Biochem Pharmacol ; 82(11): 1709-19, 2011 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-21854761

RESUMO

WAY-255348 is a potent nonsteroidal progesterone receptor (PR) antagonist previously characterized in rodents and nonhuman primates. This report describes the novel mechanism by which WAY-255348 inhibits the activity of progesterone. Most PR antagonists bind to and block PR action by inducing a unique "antagonist" conformation of the PR. However, WAY-255348 lacks the bulky side chains or chemical groups that have been associated with the conformation changes of helix 12 that lead to functional antagonism. We show that WAY-255348 achieves antagonist activity by binding to and subsequently preventing progesterone-induced nuclear accumulation, phosphorylation and promoter interactions of the PR. This effect was concentration dependent, as high concentrations of WAY-255348 alone are able to induce nuclear translocation, phosphorylation and subsequent promoter interactions resulting in partial agonist activity at these concentrations. However, at lower concentrations where nuclear accumulation and phosphorylation are prevented, the progesterone-induced DNA binding is blocked along with PR-dependent gene expression. Analysis of the PR conformation induced by WAY-255348 using a limited protease digestion assay, suggested that the WAY-255348 bound PR conformation was similar to that of a progesterone agonist-bound PR and distinct from steroidal antagonist-bound PR conformations. Furthermore, the recruitment and binding of peptides derived from nuclear receptor co-activators is consistent with WAY-255348 inducing an agonist-like conformation. Taken together, these data suggest that WAY-255348 inhibits PR action through a novel molecular mechanism that is distinct from previously studied PR modulators and may be a useful tool to further understanding of PR signaling pathways. Development of therapeutic molecules with this 'passive' antagonism mechanism may provide distinct advantages for patients with reproductive disorders or PR positive breast cancers.


Assuntos
Indóis/farmacologia , Pirróis/farmacologia , Receptores de Progesterona/antagonistas & inibidores , Transporte Ativo do Núcleo Celular , Ligação Competitiva , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Imunoprecipitação da Cromatina , Proteínas Correpressoras/metabolismo , Agonismo Parcial de Drogas , Humanos , Modelos Moleculares , Coativadores de Receptor Nuclear/metabolismo , Fosforilação , Regiões Promotoras Genéticas , Conformação Proteica , Ensaio Radioligante , Receptores de Progesterona/agonistas , Receptores de Progesterona/genética
2.
Bioorg Med Chem Lett ; 20(16): 4816-8, 2010 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-20638844

RESUMO

Non-steroidal 1-methyl-1H-pyrrole-2-carbonitrile containing tetrahydronaphthalenes and acyclic derivatives were evaluated as novel series of progesterone receptor (PR) antagonists using the T47D cell alkaline phosphatase assay. Moderate to potent PR antagonists were achieved with these scaffolds. Several compounds (e.g., 15 and 20) demonstrated low nanomolar PR antagonist potency and good selectivity versus other steroid receptors.


Assuntos
Pirróis/química , Receptores de Progesterona/antagonistas & inibidores , Tetra-Hidronaftalenos/química , Fosfatase Alcalina/metabolismo , Linhagem Celular Tumoral , Humanos , Receptores de Progesterona/metabolismo , Relação Estrutura-Atividade , Tetra-Hidronaftalenos/síntese química , Tetra-Hidronaftalenos/farmacologia
3.
Endocrinology ; 151(5): 2373-80, 2010 May.
Artigo em Inglês | MEDLINE | ID: mdl-20233794

RESUMO

Androgenetic alopecia (AGA), commonly known as male pattern baldness, is a form of hair loss that occurs in both males and females. Although the exact cause of AGA is not known, it is associated with genetic predisposition through traits related to androgen synthesis/metabolism and androgen signaling mediated by the androgen receptor (AR). Current therapies for AGA show limited efficacy and are often associated with undesirable side effects. A major hurdle to developing new therapies for AGA is the lack of small animal models to support drug discovery research. Here, we report the first rodent model of AGA. Previous work demonstrating that the interaction between androgen-bound AR and beta-catenin can inhibit Wnt signaling led us to test the hypothesis that expression of AR in hair follicle cells could interfere with hair growth in an androgen-dependent manner. Transgenic mice overexpressing human AR in the skin under control of the keratin 5 promoter were generated. Keratin 5-human AR transgenic mice exposed to high levels of 5alpha-dihydrotestosterone showed delayed hair regeneration, mimicking the AGA scalp. This effect is AR mediated, because treatment with the AR antagonist hydroxyflutamide inhibited the effect of dihydrotestosterone on hair growth. These results support the hypothesis that androgen-mediated hair loss is AR dependent and suggest that AR and beta-catenin mediate this effect. These mice can now be used to test new therapeutic agents for the treatment of AGA, accelerating the drug discovery process.


Assuntos
Alopecia/metabolismo , Modelos Animais de Doenças , Alopecia/tratamento farmacológico , Alopecia/genética , Antagonistas de Androgênios/farmacologia , Androgênios/farmacologia , Animais , Western Blotting , Linhagem Celular , Linhagem Celular Tumoral , Di-Hidrotestosterona/farmacologia , Feminino , Flutamida/análogos & derivados , Flutamida/farmacologia , Cabelo/efeitos dos fármacos , Cabelo/crescimento & desenvolvimento , Cabelo/metabolismo , Humanos , Queratina-5/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Regiões Promotoras Genéticas/genética , Ligação Proteica , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Transfecção , beta Catenina/genética , beta Catenina/metabolismo
4.
Cancer Prev Res (Phila) ; 3(3): 290-300, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20179297

RESUMO

Insulin-like growth factor-I receptor signaling contributes to the development of endometrial hyperplasia, the precursor to endometrioid-type endometrial carcinoma, in humans and in rodent models. This pathway is under both positive and negative regulation, including S6 kinase (S6K) phosphorylation of insulin receptor substrate-1 (IRS-1) at S636/639, which occurs downstream of mammalian target of rapamycin (mTOR) activation to inhibit this adapter protein. We observed activation of mTOR with a high frequency in human endometrial hyperplasia and carcinoma, but an absence of IRS-1 phosphorylation, despite high levels of activated S6K. To explore when during disease progression mammalian target of rapamycin (mTOR) activation and loss of negative feedback to IRS-1 occurred, we used the Eker rat (Tsc2(Ek/+)) model, where endometrial hyperplasia develops as a result of loss of Tsc2, a "gatekeeper" for mTOR. We observed mTOR activation early in progression in hyperplasias and in some histologically normal epithelial cells, suggesting that event(s) in addition to loss of Tsc2 were required for progression to hyperplasia. In contrast, whereas IRS-1 S636/639 phosphorylation was observed in normal epithelium, it was absent from all hyperplasias, indicating loss of IRS-1 inhibition by S6K occurred during progression to hyperplasia. Treatment with a mTOR inhibitor (WAY-129327) significantly decreased hyperplasia incidence and proliferative indices. Because progression from normal epithelium to carcinoma proceeds through endometrial hyperplasia, these data suggest a progression sequence where activation of mTOR is followed by loss of negative feedback to IRS-1 during the initial stages of development of this disease.


Assuntos
Hiperplasia Endometrial/metabolismo , Neoplasias do Endométrio/metabolismo , Endométrio/metabolismo , Proteínas Substratos do Receptor de Insulina/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Western Blotting , Hiperplasia Endometrial/patologia , Neoplasias do Endométrio/patologia , Endométrio/patologia , Feminino , Humanos , Técnicas Imunoenzimáticas , Fosforilação , Ratos , Ratos Mutantes , Transdução de Sinais , Serina-Treonina Quinases TOR , Proteína 2 do Complexo Esclerose Tuberosa , Proteínas Supressoras de Tumor/fisiologia
5.
Bioorg Med Chem Lett ; 19(23): 6666-9, 2009 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-19864132

RESUMO

Novel 5-aryl indanones, inden-1-one oximes, and inden-1-ols were synthesized and evaluated as progesterone receptor (PR) modulators using the T47D cell alkaline phosphatase assay. Both PR agonists and antagonists were achieved with appropriate 3- and 5-substitution from indanones and inden-1-ols while inden-1-one oximes provided only PR antagonists. Several compounds such as 10 and 11 demonstrated potent in vitro PR agonist potency similar to that of steroidal progesterone (1). In addition, a number of compounds (e.g., 12, 13, 17, 18) showed potent PR antagonist activity indicating the indanones and derivatives are promising PR modulator templates.


Assuntos
Indanos/farmacologia , Receptores de Progesterona/agonistas , Receptores de Progesterona/antagonistas & inibidores , Desenho de Fármacos , Indanos/síntese química , Indanos/química , Estrutura Molecular , Estereoisomerismo , Relação Estrutura-Atividade
6.
Bioorg Med Chem ; 17(22): 7802-15, 2009 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-19836247
7.
Cancer Res ; 69(15): 6171-8, 2009 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-19622772

RESUMO

Uterine leiomyomata, or fibroids, are benign tumors of the uterine myometrium that significantly affect up to 30% of reproductive-age women. Despite being the primary cause of hysterectomy in the United States, accounting for up to 200,000 procedures annually, the etiology of leiomyoma remains largely unknown. As a basis for understanding leiomyoma pathogenesis and identifying targets for pharmacotherapy, we conducted transcriptional profiling of leiomyoma and unaffected myometrium from humans and Eker rats, the best characterized preclinical model of leiomyomata. A global comparison of mRNA from leiomyoma versus myometrium in human and rat identified a highly significant overlap of dysregulated gene expression in leiomyomata. An unbiased pathway analysis using a method of gene-set enrichment based on the sigPathway algorithm detected the mammalian target of rapamycin (mTOR) pathway as one of the most highly up-regulated pathways in both human and rat tumors. To validate this pathway as a therapeutic target for uterine leiomyomata, preclinical studies were conducted in Eker rats. These rats develop uterine leiomyomata as a consequence of loss of Tsc2 function and up-regulation of mTOR signaling. Inhibition of mTOR in female Eker rats with the rapamycin analogue WAY-129327 for 2 weeks decreased mTOR signaling and cell proliferation in tumors, and treatment for 4 months significantly decreased tumor incidence, multiplicity, and size. These results identify dysregulated mTOR signaling as a component of leiomyoma etiology across species and directly show the dependence of uterine leiomyomata with activated mTOR on this signaling pathway for growth.


Assuntos
Leiomioma/metabolismo , Proteínas Quinases/metabolismo , Neoplasias Uterinas/metabolismo , Animais , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Leiomioma/genética , Miométrio/metabolismo , Miométrio/fisiologia , Análise Serial de Proteínas , Proteínas Quinases/genética , Ratos , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR , Neoplasias Uterinas/genética
8.
Endocrinology ; 150(4): 1897-903, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19022889

RESUMO

Selective estrogen receptor modulators (SERMs) are small molecules that, depending on the end point measured, may either function as estrogen receptor (ER) agonists or antagonize estrogens' agonist activity. A key feature of SERMs is the inhibition of ER agonist action on the uterus and mammary gland, but the degree of antagonism varies among compounds and end points. Bazedoxifene is a SERM that is being clinically evaluated both as a monotherapy for the prevention and treatment of osteoporosis and in combination with conjugated estrogens (CEs) for the treatment of menopausal symptoms and prevention of osteoporosis. The studies reported here compare the relative ER agonist and antagonist effects of three pharmacologically distinct SERMs (bazedoxifene, raloxifene, and lasofoxifene) on the ovariectomized mouse when administered alone or as a tissue-selective estrogen complex, a term used to describe the partnering of a SERM and one or more estrogens. At the minimum dose required to maximally reduce CE-stimulated uterine wet weight increase for each SERM, the degree of inhibition varied among the SERMs, with a rank order of bazedoxifene approximately raloxifene > lasofoxifene, in which only bazedoxifene was statistically similar to vehicle. In the mammary gland, in which amphiregulin mRNA and morphological effects were measured, bazedoxifene generally exhibited less agonist activity and was a more effective antagonist of CE than raloxifene or lasofoxifene. In summary, in an animal model evaluating estrogen-modulated uterine effects and mammary gland development, bazedoxifene exhibited less ER agonist activity than raloxifene or lasofoxifene, and, as a tissue-selective estrogen complex, bazedoxifene/CE demonstrated less mammary gland stimulation than raloxifene/CE and lasofoxifene/CE.


Assuntos
Estrogênios Conjugados (USP)/farmacologia , Glândulas Mamárias Animais/metabolismo , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Anfirregulina , Animais , Relação Dose-Resposta a Droga , Família de Proteínas EGF , Feminino , Glicoproteínas/genética , Indóis/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/genética , Glândulas Mamárias Animais/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Tamanho do Órgão/efeitos dos fármacos , Reação em Cadeia da Polimerase , Pirrolidinas/farmacologia , Cloridrato de Raloxifeno/farmacologia , Tetra-Hidronaftalenos/farmacologia , Útero/efeitos dos fármacos , Útero/metabolismo
9.
Biochem Pharmacol ; 77(2): 204-15, 2009 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-19013437

RESUMO

Progesterone receptor (PR) modulators are used in contraception and post-menopausal hormone therapy, and are under clinical development for reproductive disorders such as uterine fibroids and endometriosis. Development of tissue selective PR modulators (SPRMs) with reduced side effects and improved pharmacology represents a large unmet medical need in the area of women's health. One approach to addressing this need is to focus on the two PR isoforms PR-A and PR-B. In vitro and in vivo studies have revealed both distinct as well as overlapping gene regulation and functional responses of the two PR isoforms that suggests that PR-A selective modulators may retain a desired biological profile. We have identified a chemical series of 4-(4-chlorophenyl)-substituted piperazine carbimidothioic acid esters (PCEs) that have partial PR agonist activity and selectively activate some PR-A isoform regulated genes in T47D cells. However, full microarray analysis in these cells does not predict a global isoform selective profile for these compounds, but rather a unique gene-selective profile is observed relative to steroidal progestins. Using multiplexed peptide interaction profiling and co-activator recruitment assays we find that the mechanism of partial agonism is only partly defined by the ability to recruit known co-activators or peptides but also depends on the cell and promoter context of the gene under investigation. The data demonstrate global consequences of mechanistic and functional differences that can lead to selective biological responses of novel steroid receptor modulators.


Assuntos
Receptores de Progesterona/agonistas , Receptores de Progesterona/fisiologia , Antagonistas de Receptores de Andrógenos , Animais , Células COS , Chlorocebus aethiops , Anticoncepcionais Femininos/efeitos adversos , Anticoncepcionais Femininos/uso terapêutico , Endometriose/tratamento farmacológico , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Acetato de Medroxiprogesterona/efeitos adversos , Acetato de Medroxiprogesterona/uso terapêutico , Piperazinas/farmacologia , Reação em Cadeia da Polimerase , Progestinas/efeitos adversos , Progestinas/uso terapêutico , Receptores de Progesterona/efeitos dos fármacos , Receptores de Progesterona/genética
10.
J Steroid Biochem Mol Biol ; 112(1-3): 40-6, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18824103

RESUMO

Selective estrogen receptor modulators (SERMs) have the potential to treat estrogen sensitive diseases such as uterine leiomyoma and endometriosis, which are prevalent in reproductive age women. However, SERMs also increase the risk of developing ovarian cysts in this population, a phenomenon that is not seen in postmenopausal women. It is believed that current SERMs partially block estradiol's ability to downregulate gonadotropin-releasing hormone (GnRH) secretion from the hypothalamus thereby interfering with estradiol's negative feedback, leading to increased ovarian stimulation by gonadotropins, and cyst formation. It has been postulated that a SERM with poor brain exposure would have less negative effect on the HPO axis, therefore reducing the risk of developing ovarian cysts. In order to test this hypothesis, we identified an early marker of SERM-dependent ovarian effects: upregulation of Cyp17a1 mRNA. SERMs known to cause ovarian cysts upregulate Cyp17a1 after only 4 days of dosing and suppression of the HPO axis prevented this regulation, indicating that ovarian expression of Cyp17a1 was secondary to SERM's effect on the brain. We then characterized three SERMs with similar binding affinity and antagonist effects on the uterus for their relative brain/plasma exposure and ovarian effects. We found that the degree of brain exposure correlated very well with Cyp17a1 expression.


Assuntos
Cistos Ovarianos/metabolismo , Ovário/enzimologia , Moduladores Seletivos de Receptor Estrogênico/farmacocinética , Esteroide 17-alfa-Hidroxilase/biossíntese , Animais , Biomarcadores/metabolismo , Encéfalo/metabolismo , Relação Dose-Resposta a Droga , Receptor alfa de Estrogênio/metabolismo , Feminino , Naftalenos/administração & dosagem , Naftalenos/efeitos adversos , Naftalenos/farmacocinética , Cistos Ovarianos/patologia , Ovário/efeitos dos fármacos , Ovário/patologia , Piperidinas/administração & dosagem , Piperidinas/efeitos adversos , Piperidinas/farmacocinética , Cloridrato de Raloxifeno/administração & dosagem , Cloridrato de Raloxifeno/efeitos adversos , Cloridrato de Raloxifeno/farmacocinética , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Moduladores Seletivos de Receptor Estrogênico/administração & dosagem , Moduladores Seletivos de Receptor Estrogênico/efeitos adversos , Regulação para Cima
11.
Bioorg Med Chem Lett ; 18(18): 5015-7, 2008 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-18722119

RESUMO

A series of novel 7-(5'-cyanopyrrol-2-yl) substituted benzo[1,4]oxazepin-2-ones were prepared and tested for their progesterone receptor (PR) agonist or antagonist activity in the alkaline phosphatase assay using the human T47D breast carcinoma cell line. Both PR agonists and antagonists were achieved with an appropriate choice of 5-substitution. Several analogs were potent PR agonists (e.g., 12 and 13) or PR antagonists (e.g., 18) with good selectivity over other steroid receptors.


Assuntos
Oxazepinas/síntese química , Oxazepinas/farmacologia , Receptores de Progesterona/agonistas , Receptores de Progesterona/antagonistas & inibidores , Técnicas de Química Combinatória , Humanos , Estrutura Molecular , Oxazepinas/química , Receptores de Progesterona/metabolismo , Relação Estrutura-Atividade
12.
BMC Med Genomics ; 1: 27, 2008 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-18578861

RESUMO

BACKGROUND: Vaginal atrophy (VA) is the thinning of the vaginal epithelial lining, typically the result of lowered estrogen levels during menopause. Some of the consequences of VA include increased susceptibility to bacterial infection, pain during sexual intercourse, and vaginal burning or itching. Although estrogen treatment is highly effective, alternative therapies are also desired for women who are not candidates for post-menopausal hormone therapy (HT). The ovariectomized (OVX) rat is widely accepted as an appropriate animal model for many estrogen-dependent responses in humans; however, since reproductive biology can vary significantly between mammalian systems, this study examined how well the OVX rat recapitulates human biology. METHODS: We analyzed 19 vaginal biopsies from human subjects pre and post 3-month 17beta-estradiol treated by expression profiling. Data were compared to transcriptional profiling generated from vaginal samples obtained from ovariectomized rats treated with 17beta-estradiol for 6 hrs, 3 days or 5 days. The level of differential expression between pre- vs. post- estrogen treatment was calculated for each of the human and OVX rat datasets. Probe sets corresponding to orthologous rat and human genes were mapped to each other using NCBI Homologene. RESULTS: A positive correlation was observed between the rat and human responses to estrogen. Genes belonging to several biological pathways and GO categories were similarly differentially expressed in rat and human. A large number of the coordinately regulated biological processes are already known to be involved in human VA, such as inflammation, epithelial development, and EGF pathway activation. CONCLUSION: At the transcriptional level, there is evidence of significant overlap of the effects of estrogen treatment between the OVX rat and human VA samples.

13.
Bioorg Med Chem ; 16(13): 6589-600, 2008 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-18504132

RESUMO

Novel 7-aryl benzo[1,4]oxazepin-2-ones were synthesized and evaluated as non-steroidal progesterone receptor (PR) modulators. The structure activity relationship of 7-aryl benzo[1,4]oxazepinones was examined using the T47D cell alkaline phosphatase assay. A number of 7-aryl benzo[1,4]oxazepinones such as 10j and 10v demonstrated good in vitro potency (IC(50) of 10-30 nM) and selectivity (over 100-fold) at PR over other steroidal receptors such as glucocorticoid and androgen receptors (GR and AR). Several 7-aryl benzo[1,4]oxazepinones were active in the rat uterine decidualization model. In this in vivo model, compounds 10j and 10u were active at 3 mg/kg when dosed orally.


Assuntos
Benzazepinas/síntese química , Benzazepinas/farmacologia , Oxazepinas/síntese química , Oxazepinas/farmacologia , Receptores de Progesterona/antagonistas & inibidores , Fosfatase Alcalina/antagonistas & inibidores , Fosfatase Alcalina/metabolismo , Animais , Benzazepinas/química , Sítios de Ligação , Células COS , Chlorocebus aethiops , Feminino , Hidroxilação , Modelos Moleculares , Estrutura Molecular , Oxazepinas/química , Receptores de Progesterona/metabolismo , Relação Estrutura-Atividade
14.
Steroids ; 73(7): 689-701, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18472121

RESUMO

Progesterone receptor (PR) modulators have evolved both structurally and mechanistically over the past half-century. Classical steroidal PR agonists continue to play an important role in women's health such as in oral contraception and post-menopausal hormone therapy whereas steroid-based PR antagonists and selective PR modulators are being evaluated clinically in a wide range of gynecologic conditions. This review will focus primarily on the newer generation of PR modulators derived from structurally unique chemical scaffolds. For example, tanaproget (TNPR) is described as a non-steroidal PR agonist with high affinity and selectivity for the PR that is significantly more potent than many of its steroidal counterparts in a variety of pre-clinical efficacy models. Similarly, we present numerous examples of unique non-steroidal PR antagonists in various stages of characterization and development. A basic understanding of the structural determinants for high affinity binding of these new PR modulators to the PR ligand-binding domain (LBD) is also discussed. Finally, as the biology of the PR becomes further defined, we speculate on the future development of novel PR modulators.


Assuntos
Receptores de Progesterona , Benzoxazinas/química , Benzoxazinas/farmacologia , Estrenos/química , Estrenos/farmacologia , Feminino , Gonanos/química , Gonanos/farmacologia , Humanos , Indóis/química , Indóis/farmacologia , Oximas/química , Oximas/farmacologia , Progesterona/análogos & derivados , Progesterona/química , Progesterona/farmacologia , Isoformas de Proteínas , Receptores de Progesterona/agonistas , Receptores de Progesterona/antagonistas & inibidores , Receptores de Progesterona/química , Relação Estrutura-Atividade , Tionas/química , Tionas/farmacologia
15.
Mol Cell Endocrinol ; 287(1-2): 40-6, 2008 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-18367319

RESUMO

Selective estrogen receptor modulators (SERMs) have the unique potential to provide estrogenic effects in the skeletal and cardiovascular system, while minimizing/eliminating side effects on reproductive organs. However, despite the unifying characteristic of mixed estrogen receptor (ER) agonist/antagonist activity, compounds within this class are not interchangeable. In order to define and compare the effects of SERMs on different hormone-responsive tissues, we evaluated effects of bazedoxifene acetate (BZA), lasofoxifene (LAS) and raloxifene (RAL) in the mammary gland and uterus of the ovariectomized mouse. Endpoints measured included those regulated by estradiol alone (uterine wet weight, uterine G protein-coupled receptor 105 (GPR105) mRNA expression and mammary gland indoleamine-pyrrole 2,3 dioxygenase (INDO) mRNA expression) as well as others that required the combination of estradiol and progesterone (uterine serine protease inhibitor Kazal type 3 (Spink3) mRNA expression, mammary gland morphology and mammary gland defensin beta1 (Defbeta1) mRNA expression). The three SERMs tested had variable agonist and antagonist activity on these endpoints. In the uterus, the SERMs were mixed agonists/antagonists on estradiol-induced wet weight increase, whereas all three SERMs were estrogen receptor antagonists on GPR105 mRNA expression. However, in the presence of progesterone, BZA and RAL were agonists on Spink3 expression, while LAS was primarily an antagonist. In the mammary gland, BZA and RAL were predominantly agonists on the endpoint of mammary morphology and all three SERMs were clear agonists on Defbeta1 mRNA expression, an E+P-dependent marker. Finally, LAS and RAL had mixed agonist/antagonist activity on INDO mRNA expression, while BZA had only antagonist activity. These results demonstrate that compounds with small structural differences can elicit distinct biological responses, and that in general, SERMs tended to behave more as antagonists on endpoints requiring estrogen alone and agonists on endpoints requiring the combination of estrogen and progesterone.


Assuntos
Estradiol/farmacologia , Glândulas Mamárias Animais/efeitos dos fármacos , Progesterona/farmacologia , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Útero/efeitos dos fármacos , Animais , Biomarcadores/metabolismo , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Glicoproteínas/genética , Glicoproteínas/metabolismo , Glândulas Mamárias Animais/citologia , Camundongos , Camundongos Endogâmicos C57BL , Tamanho do Órgão/efeitos dos fármacos , Proteínas Secretadas pela Próstata/genética , Proteínas Secretadas pela Próstata/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2/metabolismo , Receptores Purinérgicos P2Y , Inibidor da Tripsina Pancreática de Kazal , Útero/citologia , beta-Defensinas/genética , beta-Defensinas/metabolismo
16.
J Med Chem ; 51(6): 1861-73, 2008 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-18318463

RESUMO

We have continued to explore the 3,3-dialkyl-5-aryloxindole series of progesterone receptor (PR) modulators looking for new agents to be used in female healthcare: contraception, fibroids, endometriosis, and certain breast cancers. Previously we reported that subtle structural changes with this and related templates produced functional switches between agonist and antagonist properties ( Fensome et al. Biorg. Med. Chem. Lett. 2002, 12, 3487; 2003, 13, 1317 ). We herein report a new functional switch within the 5-(2-oxoindolin-5-yl)-1 H-pyrrole-2-carbonitrile class of compounds. We found that the size of the 3,3-dialkyl substituent is important for controlling the functional response; thus small groups (dimethyl) afford potent PR antagonists, whereas larger groups (spirocyclohexyl) are PR agonists. The product from our optimization activities in cell-based systems and also for kinetic properties in rodents and nonhuman primates was 5-(7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1 H-indol-5-yl)-1-methyl-1 H-pyrrole-2-carbonitrile 27 (WAY-255348), which demonstrated potent and robust activity on PR antagonist and contraceptive end points in the rat and also in cynomolgus and rhesus monkeys including ovulation inhibition, menses induction, and reproductive tract morphology.


Assuntos
Desenho de Fármacos , Indóis/química , Indóis/síntese química , Indóis/farmacologia , Pirróis/química , Receptores de Progesterona/antagonistas & inibidores , Administração Oral , Fosfatase Alcalina/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Feminino , Humanos , Macaca fascicularis , Macaca mulatta , Estrutura Molecular , Ovulação/efeitos dos fármacos , Oxindóis , Pirróis/síntese química , Pirróis/farmacologia , Ratos , Receptores de Progesterona/química , Estereoisomerismo , Relação Estrutura-Atividade , Células Tumorais Cultivadas
17.
Biochem Biophys Res Commun ; 368(3): 723-8, 2008 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-18252197

RESUMO

The follicle-stimulating hormone is critical to reproductive success and is an important target for development of novel reproductive therapies. We have recently reported the development of thiazolidinone positive allosteric modulators of the follicle-stimulating hormone receptor. Here, we demonstrate that discrete modifications in the chemical structure of the thiazolidinone agonists produced compounds with different pharmacological properties. Positive allosteric modulators activated adenylate cyclase signaling (Gs). Using an ADP-ribosylation assay we found that both differing glycosylated variants of human FSH (hFSH) and selected thiazolidinone allosteric modulators of the FSHR induce activation of the Gi signaling pathway. Additionally, we observed that some analogs of this class could activate both pathways. These data suggest that the pharmacological activity of thiazolidinone modulators to the FSHR may be due to the ability of these compounds to induce association of the FSHR with either Gs or Gi signaling pathways in an analog-specific manner.


Assuntos
Células da Granulosa/metabolismo , Receptores do FSH/química , Receptores do FSH/metabolismo , Tiazolidinedionas/administração & dosagem , Tiazolidinedionas/química , Animais , Células CHO , Células Cultivadas , Cricetinae , Cricetulus , Feminino , Células da Granulosa/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptores do FSH/efeitos dos fármacos , Relação Estrutura-Atividade
18.
Maturitas ; 58(4): 366-76, 2007 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-17997058

RESUMO

BACKGROUND: Vaginal atrophy (VA) is a prevalent disorder in postmenopausal women that is characterized by decreased epithelial thickness, reduced vaginal maturation index (VMI) and increased vaginal pH. Current medical therapy consists of local or systemic replacement of estrogens. OBJECTIVE: The goal of this study was to understand, at a molecular level, the effect of estradiol (E2) on the vaginal epithelium. METHODS: Nineteen women were treated with E2 delivered through a skin patch at a dose of 0.05mg/day for 12 weeks. The diagnosis of VA was confirmed by a VMI with < or =5% superficial cells and vaginal pH>5.0. Vaginal biopsy samples were collected at baseline and after treatment. Differentially expressed mRNA transcripts in these biopsies were determined by microarray analysis. RESULTS: All 19 subjects had increased VMI (>5%) and/or reduced pH (< or =5) following treatment. Most subjects also had increased serum E2 levels and reduced serum FSH levels. Transcriptional profiling of vaginal biopsies identified over 3000 E2-regulated genes, including those involved in several key pathways known to regulate cell growth and proliferation, barrier function and pathogen defense. CONCLUSIONS: E2 controls a plethora of cellular pathways that are concordant with its profound effect on vaginal physiology. The data presented here are a useful step toward understanding the role of E2 in vaginal tissue and the development of novel therapeutics for the treatment of VA.


Assuntos
Climatério/genética , Estradiol/administração & dosagem , Regulação da Expressão Gênica/efeitos dos fármacos , Vagina/patologia , Administração Cutânea , Adulto , Idoso , Atrofia , Biópsia , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Climatério/efeitos dos fármacos , Proteínas Ricas em Prolina do Estrato Córneo , Desmogleína 1/genética , Epitélio/efeitos dos fármacos , Epitélio/patologia , Estradiol/sangue , Feminino , Hormônio Foliculoestimulante/sangue , Humanos , Concentração de Íons de Hidrogênio , Metaloproteinase 10 da Matriz/genética , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , Receptores CXCR6 , Receptores de Quimiocinas , Receptores Virais , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transcrição Gênica/efeitos dos fármacos , Vagina/efeitos dos fármacos , Vagina/metabolismo
19.
Bioorg Med Chem ; 15(20): 6556-64, 2007 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-17681796

RESUMO

We have recently discovered 5-(3-cyclopentyl-2-thioxo-2,3-dihydro-1H-benzimidazol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (14) as a potent, selective, and orally active non-steroidal progesterone receptor (PR) agonist. Compound 14 and its analog 13 possessed sub-nanomolar in vitro potency (EC(50) 0.1-0.5nM) in the T47D alkaline phosphatase assay, similar to that of the steroidal PR agonist medroxyprogesterone acetate (MPA). In contrast to MPA, 14 was highly selective (>500-fold) for the PR over both glucocorticoid and androgen receptors. In the rat uterine decidualization and complement component C3 models, 14 had oral ED(50) values of 0.02 and 0.003mg/kg, respectively, and was from 6- to 20-fold more potent than MPA. In the monkey ovulation inhibition model, compound 14 was also highly efficacious and potent with an oral ED(100) of 0.03mg/kg.


Assuntos
Nitrilas/farmacologia , Pirróis/farmacologia , Receptores de Progesterona/agonistas , Receptores de Progesterona/metabolismo , Administração Oral , Fosfatase Alcalina/metabolismo , Animais , Linhagem Celular Tumoral , Feminino , Modelos Moleculares , Estrutura Molecular , Nitrilas/química , Pirróis/química , Ratos , Ratos Sprague-Dawley , Esteroides/química , Esteroides/farmacologia
20.
J Pharmacol Exp Ther ; 323(2): 720-9, 2007 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-17673606

RESUMO

Desvenlafaxine succinate (DVS) is a recently introduced antagonist of the human norepinephrine and serotonin transporters (hNET and hSERT, respectively), currently in clinical development for use in the treatment of major depressive disorder and vasomotor symptoms associated with menopause. Initial evaluation of the pharmacological properties of DVS (J Pharmacol Exp Ther 318:657-665, 2006) revealed significantly reduced potency for the hNET expressed in membranes compared with whole cells when competing for [(3)H]nisoxetine (NIS) binding. Using hNET in transfected human embryonic kidney-293 cells, this difference in potency for DVS at sites labeled by [(3)H]NIS was found to distinguish DVS, the DVS analog rac-(1-[1-(3-chloro-phenyl)-2-(4-methylpiperazin-1-yl)-ethyl]cyclohexanol (WY-46824), methylphenidate, and the cocaine analog 3beta-(4-iodophenyl)tropane-2beta-carboxylic acid methyl ester (RTI-55) from other hNET antagonists, such as NIS, mazindol, tricyclic antidepressants, and cocaine. These differences seem not to arise from preparation-specific perturbations of ligand intrinsic affinity or antagonist-specific surface trafficking but rather from protein conformational alterations that perturb the relationships between distinct hNET binding sites. In an initial search for molecular features that differentially define antagonist binding determinants, we document that Val148 in hNET transmembrane domain 3 selectively disrupts NIS binding but not that of DVS.


Assuntos
Cicloexanóis/farmacologia , Proteínas da Membrana Plasmática de Transporte de Norepinefrina/antagonistas & inibidores , Sítios de Ligação , Ligação Competitiva , Western Blotting , Células Cultivadas , Cocaína/análogos & derivados , Cocaína/metabolismo , Succinato de Desvenlafaxina , Fluoxetina/análogos & derivados , Fluoxetina/metabolismo , Humanos , Norepinefrina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Norepinefrina/química , Ensaio Radioligante
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