RESUMO
Estradiol receptors (ER), ERalpha and ERbeta, are ligand-dependent transcription factors that regulate gene expression. Human and murine genetics suggest that ERalpha is the key target for estradiol action on bone, uterus and breast. To date, the molecular mode of action of estradiol and selective estradiol receptor modulators (SERMs) on bone is not fully understood. This is exemplified by a lack of in vitro assays that reliably predict SERM agonist activities in vivo. We hypothesized that ligand-dependent ERalpha transrepression, via protein-protein interactions at AP1, may predict estrogenic effects on bone. We modeled this using the MMP1 promoter, which encodes an AP1 binding site. We show that ICI-182780, raloxifene, 4-hydroxytamoxifen and estradiol all exhibit differential agonistic activities on the MMP1 promoter by suppressing activity by 20-80%. Transrepression efficacy and potency correlated with both uterotrophic (R(2)=0.98) and osteoprotective (R(2)=0.80) potential in the ovariectomized rat. This identifies MMP1 promoter transrepression as an agonist activity commonly shared by AF2 agonists and "antagonists" alike. Mutation analysis showed that the repression by estradiol and SERMs required correct amino acid sequences in the AF-2 domain. For instance, L540Q AF2 mutation did not alter responses to raloxifene, although it greatly increased responses to ICI-182780 (threefold) and reduced estradiol's effect by 20%. Furthermore, all tested ligands repressed the MMP1 promoter through the L540Q mutant with identical efficacy. Together, these data suggest that estradiol and SERMs share common agonist transcriptional activity via protein-protein interactions at AP1.
Assuntos
Osso e Ossos/efeitos dos fármacos , Receptor alfa de Estrogênio/agonistas , Metaloproteinase 1 da Matriz/genética , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Útero/efeitos dos fármacos , Motivos de Aminoácidos/fisiologia , Animais , Osso e Ossos/metabolismo , Regulação para Baixo/efeitos dos fármacos , Estradiol/farmacologia , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/fisiologia , Feminino , Humanos , Metaloproteinase 1 da Matriz/metabolismo , Proteínas Mutantes/fisiologia , Regiões Promotoras Genéticas/efeitos dos fármacos , Estrutura Terciária de Proteína/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Células Tumorais Cultivadas , Útero/metabolismoRESUMO
The design, synthesis, and biological activity of a series of cycloalkene acid-based niacin receptor agonists are described. This led to the discovery that tetrahydro anthranilic acid is an excellent surrogate for anthranilic acid. Several compounds were identified that were potent against the niacin receptor, had enhanced cytochrome P450 selectivity against subtypes CYP2C8 and CYP2C9, and improved oral exposure in mice.
Assuntos
Cicloexenos/síntese química , Cicloexenos/farmacologia , Ciclopentanos/síntese química , Ciclopentanos/farmacologia , Sistema Enzimático do Citocromo P-450/metabolismo , Agonistas Nicotínicos/síntese química , Agonistas Nicotínicos/farmacologia , ortoaminobenzoatos/síntese química , ortoaminobenzoatos/farmacologia , Animais , Hidrocarboneto de Aril Hidroxilases/efeitos dos fármacos , Cicloexenos/química , Cicloexenos/farmacocinética , Ciclopentanos/química , Ciclopentanos/farmacocinética , Citocromo P-450 CYP2C8 , Citocromo P-450 CYP2C9 , Desenho de Fármacos , Humanos , Camundongos , Agonistas Nicotínicos/química , Antagonistas Nicotínicos/farmacologia , ortoaminobenzoatos/química , ortoaminobenzoatos/farmacocinéticaRESUMO
Biaryl anthranilides are reported as potent and selective full agonists for the high affinity niacin receptor GPR109A. The SAR presented outlines approaches to reduce serum shift and both CYPCYP2C8 and CYP2C9 liabilities, while improving PK and maintaining excellent receptor activity. Compound 2i exhibited good in vivo antilipolytic efficacy while providing a significantly improved therapeutic index over vasodilation (flushing) with respect to niacin in the mouse model.
Assuntos
Receptores Acoplados a Proteínas G/agonistas , ortoaminobenzoatos/síntese química , Amidas/síntese química , Amidas/farmacocinética , Amidas/farmacologia , Animais , Hidrocarboneto de Aril Hidroxilases/antagonistas & inibidores , Ligação Competitiva , Células CHO , Cricetinae , Cricetulus , Citocromo P-450 CYP2C8 , Citocromo P-450 CYP2C9 , Humanos , Técnicas In Vitro , Camundongos , Microssomos Hepáticos/metabolismo , Ensaio Radioligante , Receptores Nicotínicos , Relação Estrutura-Atividade , ortoaminobenzoatos/farmacocinética , ortoaminobenzoatos/farmacologiaRESUMO
A series of androstene-3,5-diene derivatives were prepared. Despite lacking the C-3 hydroxyl previously believed necessary for ER activity, some of the analogs retained surprising affinity for ER-beta. For example, diene 4 retained excellent selectivity and potency as an ER-beta agonist and was more selective for ER-beta over the androgen receptor (AR).
Assuntos
Androstadienos/síntese química , Androstadienos/farmacologia , Receptor beta de Estrogênio/agonistas , Moduladores Seletivos de Receptor Estrogênico/síntese química , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Androstadienos/química , Animais , Sítios de Ligação/efeitos dos fármacos , Cristalografia por Raios X , Humanos , Concentração Inibidora 50 , Estrutura Molecular , Ratos , Receptores Androgênicos/efeitos dos fármacos , Moduladores Seletivos de Receptor Estrogênico/químicaRESUMO
Chemistry was developed to synthesize the title series of compounds. The ability of these novel ligands to bind to the glucocorticoid receptor was investigated. These compounds were also tested in a series of functional assays and some were found to display the profile of a dissociated glucocorticoid. The SAR of the 6,5-bicyclic series differed markedly from the previously reported 6,6-series. Molecular modeling studies were employed to understand the conformational differences between the two series of compounds, which may explain their divergent activity. Two compounds were profiled in vivo and shown to reduce inflammation in a mouse model. An active metabolite is suspected in one case.
Assuntos
Antineoplásicos/farmacologia , Compostos Bicíclicos com Pontes/química , Glucocorticoides/química , Pirazóis/química , Receptores de Glucocorticoides/efeitos dos fármacos , Animais , Antineoplásicos/síntese química , Linhagem Celular Tumoral , Humanos , Ligantes , Camundongos , Modelos Químicos , Modelos Moleculares , Receptores de Glucocorticoides/metabolismo , Relação Estrutura-AtividadeRESUMO
A series of bridged androstenediol derivatives was prepared. The bridged compounds exhibited reduced ER-beta selectivity relative to uncyclized analogs.
Assuntos
Androstenodióis/síntese química , Receptor beta de Estrogênio/antagonistas & inibidores , Moduladores Seletivos de Receptor Estrogênico/síntese química , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Androstenodióis/farmacologia , Ciclização , Receptor beta de Estrogênio/química , Receptor beta de Estrogênio/metabolismo , Humanos , Modelos Moleculares , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
A novel class of indole ligands for estrogen receptor alpha have been discovered which exhibit potent affinity and high selectivity. Substitution of the bazedoxifene skeleton to the linker present in the HTS lead 1a provided 22b which was found to be 130-fold alpha-selective and acted as an antagonist of estradiol activity in uterine tissue and MCF-7 cancer cells.
Assuntos
Receptor alfa de Estrogênio/antagonistas & inibidores , Indóis/química , Indóis/farmacocinética , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Antagonistas de Estrogênios/química , Antagonistas de Estrogênios/farmacologia , Feminino , Humanos , Concentração Inibidora 50 , Ligantes , Útero/efeitos dos fármacosRESUMO
A series of 2-9a bridged tetrahydrofluorenone derivatives were prepared which exhibited significant binding affinity for ERbeta and were highly selective.
Assuntos
Receptor beta de Estrogênio/química , Receptor beta de Estrogênio/metabolismo , Fluorenos/química , Hidrogênio/química , Fluorenos/síntese química , Humanos , Ligantes , Espectroscopia de Ressonância Magnética , Conformação MolecularRESUMO
Several tetrahydrofluorenones with a triazole fused across C7-C8 showed high levels of ERbeta-selectivity and were found to be potent ERbeta-agonists. As a class they demonstrate improved oral bioavailability in the rat over a parent class of 7-hydroxy-tetrahydrofluorenones. The most selective agonist displayed 5.7 nM affinity and 333-fold selectivity for ERbeta.
Assuntos
Compostos Azo/síntese química , Compostos Azo/farmacologia , Receptor beta de Estrogênio/agonistas , Fluorenos/química , Fluorenos/farmacologia , Animais , Compostos Azo/química , Compostos Azo/farmacocinética , Receptor beta de Estrogênio/metabolismo , Fluorenos/síntese química , Fluorenos/farmacocinética , Humanos , Ligantes , Estrutura Molecular , Ratos , Relação Estrutura-AtividadeRESUMO
A series of 6H-benzo[c]chromen-6-one and 6H-benzo[c]chromene derivatives were prepared, and the affinity and selectivity for ERalpha and ERbeta was measured. Many of the analogs were found to be potent and selective ERbeta agonists. Bis hydroxyl at positions 3 and 8 is essential for activity in a HTRF coactivator recruitment assay. Additional modifications at both phenyl rings led to compounds with ERbeta<10nM potency and >100-fold selectivity over ERalpha.
Assuntos
Derivados de Benzeno/síntese química , Benzopiranos/síntese química , Receptor beta de Estrogênio/agonistas , Derivados de Benzeno/química , Derivados de Benzeno/metabolismo , Benzopiranos/química , Benzopiranos/metabolismo , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Fluorimunoensaio , Humanos , Ligantes , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Estrogen receptor alpha (ERalpha) serine 118 (Ser118) phosphorylation modulates activation function-1 (AF1) function. Correct positioning of helix 12 promotes agonist-dependent recruitment of cyclin-dependent kinase-7 to catalyze this event. In this study we show robust cyclin-dependent kinase-7-independent, AF2 antagonist-induced Ser118 phosphorylation. Estradiol (E2) and ICI-182,780 (ICI-780) induce Ser118 phosphorylation of wild-type ERalpha and either of two helix 12 mutants, suggesting AF2-independent action, probably via shedding of 90-kDa heat shock protein. With E2 treatment, the predominantly nuclear, phosphorylated ERalpha in COS-1 cells is detergent soluble. Although levels of ICI-780-induced phosphorylation are profound, Ser118-phosphorylated ERalpha is aggregated over the nucleus or in the cytoplasm, fractionating with the cell debris and making detection in cleared lysates improbable. Selective ER modulators (SERMs) elicit a mixed response with phosphorylated ERalpha in both detergent-soluble and -insoluble compartments. Apparent ligand-induced loss of ERalpha protein from cleared lysates is thus due to ligand-induced redistribution into the pellet, not degradation. The COS-1 response to ICI-780 can be mimicked in MCF-7 cells treated with a proteasome inhibitor to block authentic ligand-induced degradation. With SERMs and antagonists, the magnitude of Ser118-phosphorylated receptor redistribution into the insoluble fraction of COS-1 cells correlates with the magnitude of authentic ERalpha degradation in MCF-7 cells. A strong inverse correlation with ligand-induced uterotropism in vivo (P < 0.0001) and direct correlation with AF2-independent transrepression of the matrix metalloprotease-1 promoter in endometrial cells in vitro are seen. These data suggest that ligand-induced Ser118 phosphorylation of ERalpha can be AF2 independent. Furthermore, they identify translocation of Ser118-phosphorylated ERalpha out of the nucleus, leading to cytoplasmic aggregation, as an antagonist pathway that may precede receptor degradation.
Assuntos
Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/metabolismo , Animais , Benzoquinonas , Células COS , Chlorocebus aethiops , Quinases Ciclina-Dependentes/efeitos dos fármacos , Quinases Ciclina-Dependentes/metabolismo , Citoplasma/efeitos dos fármacos , Citoplasma/metabolismo , Endométrio/citologia , Endométrio/efeitos dos fármacos , Estradiol/análogos & derivados , Estradiol/farmacologia , Receptor alfa de Estrogênio/agonistas , Feminino , Fulvestranto , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Proteínas de Choque Térmico HSP90/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Lactamas Macrocíclicas , Metaloproteinase 1 da Matriz/efeitos dos fármacos , Metaloproteinase 1 da Matriz/genética , Tamanho do Órgão/efeitos dos fármacos , Fosforilação , Regiões Promotoras Genéticas , Quinonas/farmacologia , Ratos , Ratos Sprague-Dawley , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Serina/metabolismo , Útero/efeitos dos fármacos , Quinase Ativadora de Quinase Dependente de CiclinaRESUMO
A series of 19-substituted androstenediol derivatives was prepared. Some of the novel analogs were surprisingly potent and selective ligands for ER-beta.
Assuntos
Androstenodiol/análogos & derivados , Androstenodiol/farmacologia , Receptor beta de Estrogênio/efeitos dos fármacos , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Androstenodiol/síntese química , Cristalografia por Raios X , Humanos , Ligantes , Modelos Moleculares , Conformação Molecular , Moduladores Seletivos de Receptor Estrogênico/síntese química , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Two novel side chains which had previously been found to enhance antagonist activity in the dihydrobenzoxathiin SERM series were applied to three existing platforms. The novel side chains did not improve the antagonist activity of the existing platforms.
Assuntos
Receptor alfa de Estrogênio/antagonistas & inibidores , Oxati-Inas/química , Moduladores Seletivos de Receptor Estrogênico/química , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Animais , Ligantes , RatosRESUMO
An optimized side chain for dihydrobenzoxathiin SERAMs was discovered and attached to four dihydrobenzoxathiin platforms. The novel SERAMs show exceptional estrogen antagonist activity in uterine tissue and an MCF-7 breast cancer cell assay.
Assuntos
Oxati-Inas/síntese química , Receptores de Estrogênio/química , Moduladores Seletivos de Receptor Estrogênico/síntese química , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Antagonistas de Estrogênios/síntese química , Antagonistas de Estrogênios/farmacologia , Feminino , Humanos , Oxati-Inas/farmacologia , Ratos , Receptores de Estrogênio/metabolismo , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Relação Estrutura-Atividade , Útero/efeitos dos fármacos , Útero/crescimento & desenvolvimentoRESUMO
A series of novel ligands for the glucocorticoid receptor containing two heterocycles were synthesized. These compounds were investigated for a dissociative profile using transrepression and transactivation assays. Several compounds were tested in vivo and showed the ability to reduce inflammation in a mouse.
Assuntos
Glucocorticoides/química , Compostos Heterocíclicos/química , Animais , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Linhagem Celular , Glucocorticoides/administração & dosagem , Glucocorticoides/farmacologia , Compostos Heterocíclicos/administração & dosagem , Compostos Heterocíclicos/farmacologia , Humanos , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/biossínteseRESUMO
Estrogens and selective estrogen receptor modulators (SERMs) are prescribed widely in the clinic to alleviate symptoms in postmenopausal women, and they are metabolized to reactive intermediates, which may elicit adverse effects. As part of our efforts to develop safer SERMs, in vitro covalent protein binding of (2S,3R)-(+)-3-(4-hydroxyphenyl)-2-[4-(2-piperidin-1-ylethoxy)phenyl]-2,3-dihydro-1,4-benzoxathiin-6-ol (I) was evaluated. Radioactivity from [3H]I became covalently bound to proteins in a fashion that was both time- and NADPH-dependent in human liver microsomes and reached a value of 1106 pmol equiv/mg protein following a 45 min incubation. At least three pathways are involved in the bioactivation of I, namely, oxidative cleavage of the dihydrobenzoxathiin moiety to give a hydroquinone/para-benzoquinone redox couple, hydroxylation at position 5 or 7 of the benzoxathiin moiety leading to an o-quinone intermediate, and metabolism of the piperidine ring to give an iminium ion. The latter reactive intermediate was identified as its bis-cyano adduct when human liver microsomal incubations were performed in the presence of sodium cyanide. Structural modification of I, including a replacement of the piperidine with a pyrrolidine group, led to (2S,3R)-(+)-3-(3-hydroxyphenyl)-2-[4-(2-pyrrolidin-1-ylethoxy)phenyl]-2,3-dihydro-1,4-benzoxathiin-6-ol (II), which did not form a reactive iminium ion. Following the incubation of II with human liver microsomes, covalent binding to proteins was reduced (461 pmol equiv/mg protein), the residual level of binding apparently due to the formation of a rearranged biphenyl quinone type metabolite. Studies with inhibitory antibodies and chemical inhibitors showed that P450 3A4 was the primary enzyme responsible for oxidative bioactivation of I and II in human liver microsomes. These studies thus demonstrated that gaining an understanding of bioactivation mechanisms may be exploited in terms of guiding structural modifications of drug candidates to minimize covalent protein binding and, hopefully, to lower the potential for drug-mediated adverse effects.
Assuntos
Sistema Enzimático do Citocromo P-450/fisiologia , Microssomos Hepáticos/metabolismo , Oxati-Inas/farmacocinética , Moduladores Seletivos de Receptor Estrogênico/farmacocinética , Benzoquinonas/metabolismo , Biotransformação , Citocromo P-450 CYP3A , Hepatócitos/metabolismo , Humanos , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Ligação ProteicaRESUMO
The discovery, synthesis, and SAR of chromanes as ER alpha subtype selective ligands are described. X-ray studies revealed that the origin of the ER alpha-selectivity resulted from a C-4 trans methyl substitution to the cis-2,3-diphenyl-chromane platform. Selected compounds from this class demonstrated very potent in vivo antagonism of estradiol in an immature rat uterine weight assay, effectively inhibited ovariectomy-induced bone resorption in a 42 days treatment paradigm, and lowered serum cholesterol levels in ovx'd adult rat models. The best antagonists 8F and 12F also exhibited potent inhibition of MCF-7 cell growth and were shown to be estrogen receptor down-regulators (SERDs).
Assuntos
Cromanos/química , Cromanos/farmacologia , Receptor alfa de Estrogênio/metabolismo , Moduladores Seletivos de Receptor Estrogênico/química , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Animais , Sítios de Ligação , Linhagem Celular , Feminino , Expressão Gênica/efeitos dos fármacos , Humanos , Ligantes , Modelos Químicos , Estrutura Molecular , Tamanho do Órgão , Ligação Proteica , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Útero/efeitos dos fármacosRESUMO
[reaction: see text] During the course of drug metabolism studies, a major metabolite of compound 1 was detected in rhesus monkeys and assigned structure 4. The intriguing biotransformation of 1 leading to 4 was confirmed by a 19-step total synthesis starting from resorcinol (11), the key feature of which was the construction of the oxygen bridge utilizing a phenolic oxidation and trapping sequence. In addition, the synthesis of a related metabolite (5) is described.
Assuntos
Receptor alfa de Estrogênio/antagonistas & inibidores , Osteoporose/tratamento farmacológico , Oxati-Inas/síntese química , Oxati-Inas/farmacologia , Moduladores Seletivos de Receptor Estrogênico/síntese química , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Animais , Biotransformação , Macaca mulatta , Modelos Moleculares , Conformação Molecular , Oxirredução , Oxigênio/química , Resorcinóis/metabolismoRESUMO
The ring oxygen and sulfur analogs of lasofoxifene, 1a and 1b, were synthesized in an attempt to impart ERalpha selectivity, as found in the closely related dihydrobenzoxathiin compound I, recently discovered in these laboratories. The resulting isochroman and isothiochroman compounds were found to exhibit equipotent binding affinities to the ER isoforms and were less active in the inhibition of estradiol-triggered uterine growth when compared to I and lasofoxifene.
Assuntos
Cromanos/síntese química , Cromanos/farmacologia , Receptor alfa de Estrogênio/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Estradiol/farmacologia , Feminino , Humanos , Concentração Inibidora 50 , Ligantes , Modelos Moleculares , Ligação Proteica , Isoformas de Proteínas , Relação Estrutura-Atividade , Útero/efeitos dos fármacos , Útero/crescimento & desenvolvimentoRESUMO
A series of dihydrobenzoxathiin SERAMs with alkylated pyrrolidine side chains or alkylated linkers was prepared. Minor modifications in the side chain or linker resulted in significant effects on biological activity, especially in uterine tissue.